Tumor-infiltrating Neutrophils Research Articles

Methionine Restriction Reduces Lung Cancer Progression and Increases Chemotherapy Response.

Targeting tumor metabolism through dietary interventions is an area of growing interest, and may help to improve the significant mortality of aggressive cancers, including non-small cell lung cancer (NSCLC). Here we show that the restriction of methionine in the aggressive KRAS /Lkb1- mutant NSCLC autochthonous mouse model drives decreased tumor progression and increased carboplatin treatment efficacy. Importantly, methionine restriction during early stages of tumorigenesis prevents the lineage switching known to occur in the model, and alters the tumor immune microenvironment (TIME) to have fewer tumor-infiltrating neutrophils. Mechanistically, mutations in LKB1 are linked to anti-oxidant production through changes to cystathionine-β-synthase (CBS) expression. Human cell lines with rescued LKB1 show increased CBS levels and resistance to carboplatin, which can be partially rescued by methionine restriction. Furthermore, LKB1 rescued cells, but not mutant cells, show less G2- M arrest and apoptosis in high methionine conditions. Knock-down of CBS sensitized both LKB1 mutant and non-mutated lines to carboplatin, again rescuing the carboplatin resistance of the LKB1 rescued lines. Given that immunotherapy is commonly combined with chemotherapy for NSCLC, we next wanted to understand if T cells are impaired by MR. Therefore, we examined the ability of T cells from MR and control tumor bearing mice to proliferate in culture and found that T cells from MR treated mice had no defects in proliferation, even though we continued the MR conditions ex vivo . We also identified that CBS is most highly correlated with smoking, adenocarcinomas with alveolar and bronchiolar features, and adenosquamous cell carcinomas, implicating its roles in oxidative stress response and lineage fate in human tumors. Taken together, we have shown the importance of MR as a dietary intervention to slow tumor growth and improve treatment outcomes for NSCLC.

SCARB2 associates with tumor-infiltrating neutrophils and predicts poor prognosis in breast cancer.

The tumor microenvironment (TME) plays a crucial role in various aspects of breast cancer development and metastasis. Nevertheless, the expression, prognostic significance, and correlation with clinical features of SCARB2 in breast cancer, as well as the infiltrative characteristics of TME, remain largely unknown. We analyzed the differential presentation of SCARB2 mRNA in breast cancer tissues and nontumorous breast tissues and prognosis by The Cancer Genome Atlas (TCGA) and the Genotype-Tissue Expression (GTEx) databases. Additionally, the Tumor Immunity Estimation Resource (TIMER) was taken to evaluate the correlation between SCARB2 mRNA presence and tumor-infiltrating immune cells and immune checkpoints in the TME in breast cancer. We performed multiple immunohistochemical staining to verify the SCARB2 protein expression in breast cancer tissues and its relationship to immune cells and checkpoints and clinicopathological features. We identified elevated SCARB2 expression in breast cancer tissues, and high SCARB2 protein presentation was associated with advanced clinical stage and unfavorable prognosis. In addition, enhanced SCARB2 protein presence was closely correlated with up-regulation CD66b+ neutrophils infiltration in tumor tissues (r = 0.210, P < 0.05) and CD68 + CD163+ M2 macrophages in the interstitium (r = 0.233, P < 0.05), as well as the immune checkpoints, including PD-1 (r = 0.314, P < 0.01) protein expression. SCARB2 holds promise for predicting the clinical outcome of breast cancer patients and could serve as a potential therapeutic target.

β-Lapachone promotes the recruitment and polarization of tumor-associated neutrophils (TANs) toward an antitumor (N1) phenotype in NQO1-positive cancers

ABSTRACT NAD(P)H:quinone oxidoreductase 1 (NQO1) is overexpressed in most solid cancers, emerging as a promising target for tumor-selective killing. β-Lapachone (β-Lap), an NQO1 bioactivatable drug, exhibits significant antitumor effects on NQO1-positive cancer cells by inducing immunogenic cell death (ICD) and enhancing tumor immunogenicity. However, the interaction between β-Lap-mediated antitumor immune responses and neutrophils, novel antigen-presenting cells (APCs), remains unknown. This study demonstrates that β-Lap selectively kills NQO1-positive murine tumor cells by significantly increasing intracellular ROS formation and inducing DNA double strand breaks (DSBs), resulting in DNA damage. Treatment with β-Lap efficiently eradicates immunocompetent murine tumors and significantly increases the infiltration of tumor-associated neutrophils (TANs) into the tumor microenvironment (TME), which plays a crucial role in the drug’s therapeutic efficacy. Further, the presence of β-Lap-induced antigen medium leads bone marrow-derived neutrophils (BMNs) to directly kill murine tumor cells, aiding in dendritic cells (DCs) recruitment and significantly enhancing CD8+ T cell proliferation. β-Lap treatment also drives the polarization of TANs toward an antitumor N1 phenotype, characterized by elevated IFN-β expression and reduced TGF-β cytokine expression, along with increased CD95 and CD54 surface markers. β-Lap treatment also induces N1 TAN-mediated T cell cross-priming. The HMGB1/TLR4/MyD88 signaling cascade influences neutrophil infiltration into β-Lap-treated tumors. Blocking this cascade or depleting neutrophil infiltration abolishes the antigen-specific T cell response induced by β-Lap treatment. Overall, this study provides comprehensive insights into the role of tumor-infiltrating neutrophils in the β-Lap-induced antitumor activity against NQO1-positive murine tumors.

TET2-STAT3-CXCL5 nexus promotes neutrophil lipid transfer to fuel lung adeno-to-squamous transition.

Phenotypic plasticity is a rising cancer hallmark, and lung adeno-to-squamous transition (AST) triggered by LKB1 inactivation is significantly associated with drug resistance. Mechanistic insights into AST are urgently needed to identify therapeutic vulnerability in LKB1-deficient lung cancer. Here, we find that ten-eleven translocation (TET)-mediated DNA demethylation is elevated during AST in KrasLSL-G12D/+; Lkb1L/L (KL) mice, and knockout of individual Tet genes reveals that Tet2 is required for squamous transition. TET2 promotes neutrophil infiltration through STAT3-mediated CXCL5 expression. Targeting the STAT3-CXCL5 nexus effectively inhibits squamous transition through reducing neutrophil infiltration. Interestingly, tumor-infiltrating neutrophils are laden with triglycerides and can transfer the lipid to tumor cells to promote cell proliferation and squamous transition. Pharmacological inhibition of macropinocytosis dramatically inhibits neutrophil-to-cancer cell lipid transfer and blocks squamous transition. These data uncover an epigenetic mechanism orchestrating phenotypic plasticity through regulating immune microenvironment and metabolic communication, and identify therapeutic strategies to inhibit AST.

Tumor-Intrinsic Enhancer of Zeste Homolog 2 Controls Immune Cell Infiltration, Tumor Growth, and Lung Metastasis in a Triple-Negative Breast Cancer Model.

Triple-negative breast cancer (TNBC) is an aggressive and highly metastatic type of tumor. TNBC is often enriched in tumor-infiltrating neutrophils (TINs), which support cancer growth in part by counteracting tumor-infiltrating lymphocytes (TILs). Prior studies identified the enhancer of zeste homolog 2 (EZH2) as a pro-tumor methyltransferase in primary and metastatic TNBCs. We hypothesized that EZH2 inhibition in TNBC cells per se would exert antitumor activity by altering the tumor immune microenvironment. To test this hypothesis, we used CRISPR to generate EZH2 gene knockout (KO) and overexpressing (OE) lines from parent (wild-type-WT) 4T1 cells, an established murine TNBC model, resulting in EZH2 protein KO and OE, respectively. In vitro, EZH2 KO and OE cells showed early, transient changes in replicative capacity and invasiveness, and marked changes in surface marker profile and cytokine/chemokine secretion compared to WT cells. In vivo, EZH2 KO cells showed significantly reduced primary tumor growth and a 10-fold decrease in lung metastasis compared to WT cells, while EZH2 OE cells were unchanged. Compared to WT tumors, TIN:TIL ratios were greatly reduced in EZH2 KO tumors but unchanged in EZH2 OE tumors. Thus, EZH2 is key to 4T1 aggressiveness as its tumor-intrinsic knockout alters their in vitro secretome and in vivo primary tumor growth, TIN/TIL poise, and metastasis.

A pan-cancer analysis of the microbiome in metastatic cancer

Microbial communities are resident to multiple niches of the human body and are important modulators of the host immune system and responses to anticancer therapies. Recent studies have shown that complex microbial communities are present within primary tumors. To investigate the presence and relevance of the microbiome in metastases, we integrated mapping and assembly-based metagenomics, genomics, transcriptomics, and clinical data of 4,160 metastatic tumor biopsies. We identified organ-specific tropisms of microbes, enrichments of anaerobic bacteria in hypoxic tumors, associations between microbial diversity and tumor-infiltrating neutrophils, and the association of Fusobacterium with resistance to immune checkpoint blockade (ICB) in lung cancer. Furthermore, longitudinal tumor sampling revealed temporal evolution of the microbial communities and identified bacteria depleted upon ICB. Together, we generated a pan-cancer resource of the metastatic tumor microbiome that may contribute to advancing treatment strategies.

Abstract 2680: Knockout on enhancer of zeste homolog 2 in triple-negative breast cancer model alters immune signaling and decreases tumor progression and lung metastasis

Abstract Triple-negative breast cancer (TNBC) is an aggressive and highly metastatic subtype of breast cancer. Lack of known targets, patient-to-patient variations of target antigens and resistance to immune checkpoint inhibitors make TNBC one of the most challenging cancers to treat. TNBC is enriched in tumor-infiltrating neutrophils (TINs). TINs are key contributors to pro-tumorigenic processes linked to “immunologically cold” tumors, in part by preventing recruitment of T-lymphocytes (TILs). Prior studies identified the Enhancer of zeste homolog 2 (EZH2) as a prominent methyltransferase expressed in primary and metastatic TNBC cells. EZH2 is an upstream regulator of the stimulator of interferon genes (STING), a critical innate sensor mediating anti-tumor responses that is inhibited in cold tumors. To date, EZH2/STING signaling in TINs is relatively unknown. Here, we hypothesized that EZH2 inhibition and subsequent STING activation could exert anti-tumor activity by modulating TINs. We derived EZH2-knockout (KO) and EZH2-overexpressed (OX) cell lines from the well-established 4T1 TNBC mouse cell line. We demonstrate a 10-fold decrease in lung metastasis and significant reduction of 4T1 EZH2 KO tumor growth compared to the parent 4T1 cell line. Concomitantly, the TIN:TIL ratio was significantly reduced in 4T1 EZH2 KO vs. control tumors. Overexpression of EZH2 in 4T1 TNBC cells did not affect the primary tumor growth or lung metastasis. A therapeutic study using the repeated injection of the STING agonist MSA-2 after tumor engraftment in vivo also demonstrated a decrease in lung metastasis and tumor growth in MSA-2 vs. vehicle injection with both 4T1 parent and EZH2 KO cell lines. Together, our findings suggest a key role for STING pathway modulation and TIN/TIL poise in primary TNBC progression and lung metastasis. Citation Format: Lenore Monterroza, Maria Parrilla, Sarah Samaranayake, Dormarie Rivera, Brian Dobosh, Periasamy Selvaraj, Rabindra M. Tirouvanziam. Knockout on enhancer of zeste homolog 2 in triple-negative breast cancer model alters immune signaling and decreases tumor progression and lung metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2680.

The relationship between tumor-infiltrating neutrophils and clinical outcomes in patients with resectable hepatocellular carcinoma

BackgroundThe impact of tumor-infiltrating neutrophils (TINs) on clinical outcomes has been reported in various cancer types, but their role in hepatocellular carcinoma (HCC) has not been fully evaluated. The aim of this study was to investigate the prognostic values for TINs in HCC patients undergoing curative resection.MethodsWe assessed immune markers (CD3, CD4, CD8, CD66b) using immunohistochemistry in 115 patients who underwent curative resection for HCC. We analyzed the prognostic values for tumor-infiltrating immune cells, including neutrophils, and other clinicopathological factors.ResultsIn the Multivariate Cox analysis of overall survival (OS), alpha-fetoprotein (AFP) ≥ 100 ng/mL (hazard ratio (HR), 2.74, 95% confidence interval (CI), 1.17–6.44; P = 0.021) and Barcelona Clinic Liver Cancer (BCLC) B/C stage (HR, 3.98, 95% CI, 1.68–9.43; P = 0.020) were found to be independent poor prognostic factors in HCC patients undergoing resection. The presence of CD66b+TINs was observed in 66 (57.4%) patients. However, CD66b+TINs were not associated with recurrence-free survival and OS.ConclusionsOur study identified low CD66b+TINs in resectable HCC, and CD66b+ TINs did not have a significant role for the clinical outcomes of patients undergoing curative resection. The results suggest that TINs may play a role in more advanced stages of HCC.

Increased Apolipoprotein A1 Expression Correlates with Tumor-Associated Neutrophils and T Lymphocytes in Upper Tract Urothelial Carcinoma.

An increased neutrophil-to-lymphocyte ratio (NLR) is a poor prognostic biomarker in various types of cancer, because it reflects the inhibition of lymphocytes in the circulation and tumors. In urologic cancers, upper tract urothelial carcinoma (UTUC) is known for its aggressive features and lack of T cell infiltration; however, the association between neutrophils and suppressed T lymphocytes in UTUC is largely unknown. In this study, we examined the relationship between UTUC-derived factors and tumor-associated neutrophils or T lymphocytes. The culture supernatant from UTUC tumor tissue modulated neutrophils to inhibit T cell proliferation. Among the dominant factors secreted by UTUC tumor tissue, apolipoprotein A1 (Apo-A1) exhibited a positive correlation with NLR. Moreover, tumor-infiltrating neutrophils were inversely correlated with tumor-infiltrating T cells. Elevated Apo-A1 levels in UTUC were also inversely associated with the population of tumor-infiltrating T cells. Our findings indicate that elevated Apo-A1 expression in UTUC correlates with tumor-associated neutrophils and T cells. This suggests a potential immunomodulatory effect on neutrophils and T cells within the tumor microenvironment, which may represent therapeutic targets for UTUC treatment.

Early Contrast-Enhanced MR for Diagnosing Complete Tumor Response of Locally Advanced Esophageal Squamous Cell Carcinoma After Neoadjuvant Therapy: A Retrospective Comparative Study.

This study assessed the performance of early contrast-enhanced magnetic resonance (ECE-MR) in the detecting of complete tumor response (ypT0) in patients with esophageal squamous cell carcinoma following neoadjuvant therapy. Preoperative MR images of consecutive patients who underwent neoadjuvant therapy and surgical resection were reviewed retrospectively. The accuracy of ECE-MR and T2WI+DWI was evaluated by comparing the findings with pathological results. Receiver operating characteristic curve analysis was used to assess the diagnostic performance, and DeLong method was applied to compare the areas under the curves (AUC). Chi-squared analysis was conducted to explore the difference in pathological changes. A total of 198 patients (mean age 62.6 ± 7.8 years, 166 men) with 201 lesions were included. The AUC of ECE-MR was 0.85 (95% CI 0.79-0.90) for diagnosing ypT1-4, which was significantly higher than that of T2WI+DWI (AUC 0.69, 95% CI 0.63-0.76, p < 0.001). The diagnostic performance of both T2WI+DWI and ECE-MR improved with increasing tumor stage. The AUCs of ECE-MRI were higher in ypT1 and ypT2 tumors than T2WI+DWI. Degree 2-3 tumor-infiltrating lymphocytes and neutrophils were commonly seen in ypT0 tumors misdiagnosed by ECE-MR. Visual evaluation of ECE-MR is a promising diagnostic protocol for the detection of complete tumor response, especially for differentiation with early stage tumors. The accurate diagnosis of complete tumor response after neoadjuvant therapy using imaging modalities is of important significance for clinical decision-making for patients with esophageal squamous cell carcinoma. It is hoped that early contrast-enhanced MR will provide supportive advice for the development of individualized treatment options for patients.

78 Targeting tumor-promoting myeloid cell functions in head & neck cancer

Myeloid cells are a major component of immune cells infiltrating tumors, interacting with each other, with tumor cells and other stromal cells, and demonstrating a prominent plasticity. Tumor-associated myeloid cells, such as neutrophils (or so-called MDSCs: suppressory neutrophils) contribute to cancer progression via multiple mechanisms. Neutrophils are the most abundant myeloid blood cells, which have been shown to exert heterogeneous phenotypic and functional states: reaching from anti- to protumoral. On the one hand these cells can suppress anti-tumor immune responses, support tumor angiogenesis or metastatic dissemination. On the other hand, neutrophils actively kill tumor cells, or activate T-cell dependent immune responses.There is still little information on tumor-infiltrating neutrophils and their clinical relevance. Nevertheless, neutrophils were demonstrated to be associated with poor clinical outcome in patients with different kinds of cancer, including renal cancer, non-small-cell lung carcinoma, melanoma and head and neck cancer. Enrichment of neutrophils was also associated with metastases in various tumor entities. The recruitment of neutrophils within the tumor microenvironment is strictly regulated by chemokines, such as CXCL2 or CXCL8/IL-8.Neutrophil activity strongly depends on the stimulus present in the microenvironment. Since these cells are highly plastic and can easily alter their functionality, they are an ideal target for designing anti-tumor therapies. However, most of therapeutic strategies to target neutrophils in tumor-bearing hosts are dealing with the blocking of their migration into the tumor site. The efficiency of such approaches is, however, limited, as it does not address heterogeneity of these cells, and thus depletes as well antitumoral cells, leading to cancer progression and infectious complications.Given a critical role of neutrophils in tumor progression, we tested several alternative strategies to neutralize pro-tumoral activity of these cells, including reprogramming and inhibition of their immunosuppressive functions. Our recent data using own designed CCL2 peptide MyeloMIB show promising therapeutic results in head and neck cancer preclinical models. Application of this peptide leads not only to the specific inhibition of neutrophil migration into tumor tissue, but also reprograms these cells into anti-tumoral state. These results suggest a dual-edged role of neutrophils as essential regulators of anti-cancer immune responses and argue for approaches fostering anti-cancer activity of these cells, not only their migration, during cancer immunotherapy.

Intratumoral CXCR4hi neutrophils display ferroptotic and immunosuppressive signatures in hepatoblastoma.

Pediatric hepatoblastoma (HB) is the most common primary liver malignancy in infants and children. With great diversity and plasticity, tumor-infiltrating neutrophils were one of the most determining factors for poor prognosis in many malignant tumors. In this study, through bulk RNA sequencing for sorted blood and tumor-infiltrated neutrophils and comparison of neutrophils in tumor and para-tumor tissue by single-cell sequencing, we found that intratumoral neutrophils were composed of heterogenous functional populations at different development stages. Our study showed that terminally differentiated neutrophils with active ferroptosis prevailed in tumor tissue, whereas, in para-tumor, pre-fate naïve neutrophils were dominant and ferroptotic neutrophils dispersed in a broad spectrum of cell maturation. Gene profiling and in vitro T-cell coculture experiment confirmed that one of main functional intratumoral neutrophils was mainly immunosuppressive, which relied on the activation of ferroptosis. Combining the bulk RNA-seq, scRNA-seq data, and immunochemistry staining of tumor samples, CXCL12/CXCR4 chemotaxis pathway was suggested to mediate the migration of neutrophils in tumors as CXCR4 highly expressed by intratumoral neutrophils and its ligand CXCL12 expressed much higher level in tumor than that in para-tumor. Moreover, our study pinpointed that infiltrated CXCR4hi neutrophils, regardless of their differential distribution of cell maturation status in HB tumor and para-tumor regions, were the genuine perpetrators for immune suppression. Our data characterized the ferroptosis-dependent immunosuppression energized by intratumoral CXCR4 expression neutrophils and suggest a potential cell target for cancer immunotherapies.

Systemic and local immunosuppression in glioblastoma and its prognostic significance.

The effectiveness of tumor therapy, especially immunotherapy and oncolytic virotherapy, critically depends on the activity of the host immune cells. However, various local and systemic mechanisms of immunosuppression operate in cancer patients. Tumor-associated immunosuppression involves deregulation of many components of immunity, including a decrease in the number of T lymphocytes (lymphopenia), an increase in the levels or ratios of circulating and tumor-infiltrating immunosuppressive subsets [e.g., macrophages, microglia, myeloid-derived suppressor cells (MDSCs), and regulatory T cells (Tregs)], as well as defective functions of subsets of antigen-presenting, helper and effector immune cell due to altered expression of various soluble and membrane proteins (receptors, costimulatory molecules, and cytokines). In this review, we specifically focus on data from patients with glioblastoma/glioma before standard chemoradiotherapy. We discuss glioblastoma-related immunosuppression at baseline and the prognostic significance of different subsets of circulating and tumor-infiltrating immune cells (lymphocytes, CD4+ and CD8+ T cells, Tregs, natural killer (NK) cells, neutrophils, macrophages, MDSCs, and dendritic cells), including neutrophil-to-lymphocyte ratio (NLR), focus on the immune landscape and prognostic significance of isocitrate dehydrogenase (IDH)-mutant gliomas, proneural, classical and mesenchymal molecular subtypes, and highlight the features of immune surveillance in the brain. All attempts to identify a reliable prognostic immune marker in glioblastoma tissue have led to contradictory results, which can be explained, among other things, by the unprecedented level of spatial heterogeneity of the immune infiltrate and the significant phenotypic diversity and (dys)functional states of immune subpopulations. High NLR is one of the most repeatedly confirmed independent prognostic factors for shorter overall survival in patients with glioblastoma and carcinoma, and its combination with other markers of the immune response or systemic inflammation significantly improves the accuracy of prediction; however, more prospective studies are needed to confirm the prognostic/predictive power of NLR. We call for the inclusion of dynamic assessment of NLR and other blood inflammatory markers (e.g., absolute/total lymphocyte count, platelet-to-lymphocyte ratio, lymphocyte-to-monocyte ratio, systemic immune-inflammation index, and systemic immune response index) in all neuro-oncology studies for rigorous evaluation and comparison of their individual and combinatorial prognostic/predictive significance and relative superiority.

Prognostic significance and immune escape implication of tumor-infiltrating neutrophil plasticity in human head and neck squamous cell carcinoma.

Tumor-infiltrating neutrophils play a crucial role in the progression of head and neck squamous cell carcinoma (HNSCC). Here, we aimed to statistically quantify the plasticity of HNSCC-infiltrating N2/N1 neutrophils and examine its impacts on survival and immune infiltration landscape. A retrospective study of 80 patients who underwent curative surgical resection for HNSCC between 2014 and 2017 was conducted in this study. HNSCC-infiltrating neutrophil phenotypes were classified using immunofluorescence staining, and the N2/N1 neutrophil plasticity was evaluated via the ratio of N2/N1 neutrophils. We then assessed the correlations between N2/N1 neutrophil plasticity, clinicopathological characteristics, and immune infiltration landscape using rigorous statistical methods. Infiltration variations of N1 and N2 neutrophils were observed between the tumor nest (TN) and tumor stroma (TS), with TN exhibiting higher N2 neutrophil infiltration and lower N1 neutrophil infiltration. High ratios of N2/N1 neutrophils were correlated with advanced TNM stage, large tumor size and invasion of adjacent tissue. High infiltration of N2 neutrophils was associated with decreased overall and relapse-free survival, which were opposite for N1 neutrophils. The independent prognostic role of N2/N1 neutrophil plasticity, particularly within the TN region, was confirmed by multivariate analyses. Moreover, the ratio of N2/N1 neutrophils within the TN region showed correlations with high CD8+ T cells infiltration and low FOXP3+ Tregs infiltration. We identify HNSCC-infiltrating N2/N1 neutrophil plasticity as a crucial prognostic indictor which potentially reflects the tumor microenvironment (TME) and immune escape landscape within HNSCC tissues. Further investigations and validations may provide novel therapeutic strategies for personalized immunomodulation in HNSCC patients.

Neutrophils in HNSCC Can Be Associated with Both a Worse or Favorable Prognosis.

The prognostic significance of tumor-infiltrating neutrophils in head and neck squamous cell carcinoma (HNSCC) is poorly understood. It is unclear how the presence of neutrophils affects prognosis due to their polarization into cytotoxic N1 or immunosuppressive N2. Therefore, we determined the number of CD66b+ neutrophil granulocytes separately in the stromal and epithelial compartments in cancer tissues from 397 patients with HNSCC. Tumor samples from six historical patient groups were processed into tissue microarrays and stained immunohistochemically. In total, 21.9% were HPV positive (p16+). Neutrophil counts were much lower in the stromal compartment (372 ± 812) than in the epithelial cancer compartment (1040 ± 1477) (p < 0.001), with large differences between groups. In three groups with high neutrophil infiltration, high rates were associated with a favorable prognosis, whereas in two groups, high rates were a negative prognostic factor. In p16- oropharyngeal and hypopharyngeal cancer high infiltration was associated with a favorable prognosis. Cancers with an exclusion of neutrophils in the epithelial compartment were associated with improved prognosis. In oropharyngeal and hypopharyngeal HPV-negative cancer high neutrophil infiltration rates were clearly associated with prolonged survival. Neutrophil granulocytes in HNSCC may contribute to a favorable or unfavorable prognosis.

Immature stroma and high infiltration of CD15+ cells are predictive markers of poor prognosis in different subsets of patients with pancreatic cancer.

Preoperative treatment is commonly carried out for borderline resectable pancreatic ductal adenocarcinoma (PDAC). However, the relationship between the combination of immune cells in the tumor microenvironment and their intratumoral heterogeneity along with their association with histological findings remains unclear, especially in patients receiving preoperative chemotherapy. We aimed to explore the therapeutic strategies for patients with PDAC with poor prognosis after receiving chemotherapy based on histological and immunological microenvironmental classifications. We investigated the correlation between the prognosis and histological immune microenvironmental factors of patients who initially underwent surgery (n = 100) and were receiving gemcitabine plus nab-paclitaxel (GEM + nabPTX) as preoperative chemotherapy (n = 103). Immune profiles were generated based on immune cell infiltration into the tumor, and their correlation with patient outcomes and histological features was analyzed. Tumor-infiltrating neutrophils (TINs) were identified as independent poor prognostic factors using multivariate analysis in both surgery-first and preoperative chemotherapy groups. The patients were further classified into four groups based on immune cell infiltration into the tumor. Patients with high CD15 infiltration into the tumor and immature stroma at the cancer margins showed the worst prognosis in the preoperative chemotherapy group. The analysis of mRNA expression and immunohistochemical features revealed that CXCR2, the receptor for CXCL8, was correlated with disease-free and overall survival. We inferred that patients with immature stroma at the margins and high infiltration of CD15+ neutrophils within the tumor showed the worst prognosis and they could particularly benefit from treatment with inhibitors targeting CXCR2 or CXCL8.

Prognostic significance of LRPPRC and its association with immune infiltration in liver hepatocellular carcinoma.

Leucine rich pentatricopeptide repeat containing (LRPPRC) protein is a multifunctional protein involved in cell cycle progression and tumor development. However, its prognostic significance and association with immune infiltration in Liver hepatocellular carcinoma (LIHC) remain unclear. We utilized transcriptomic and clinical data from The Cancer Genome Atlas (TCGA) and Genotype Tissue Expression (GTEx) databases of LIHC patients to investigate the potential pro-cancer role of LRPPRC, including differential expression of LRPPRC in LIHC, prognostic value, clinicopathological features, immune cell infiltration relevance and function enrichment analysis. Our findings suggest that LRPPRC is upregulated in LIHC and exhibits correlations with survival, clinical stage, and tumor grade in LIHC patients. Additionally, immune infiltration analysis revealed significant negative correlations between LRPPRC expression and multiple tumor-infiltrating immune cells, including CTLs, DCs, pDCs, B cells, Th17 cells, neutrophils, T cells, Mast cells, Th1 cells, Tregs, and NK cells, whereas a significant positive correlation was observed with infiltration of Th2 cells, T helper cells and Tcms. Furthermore, functional enrichment analysis indicated that LRPPRC may be involved in G2m checkpoint, mitotic spindle, E2f targets, Wnt Beta catenin signaling, spermatogenesis and other processes.

Bcl-xL targeting eliminates ageing tumor-promoting neutrophils and inhibits lung tumor growth

Elevated peripheral blood and tumor-infiltrating neutrophils are often associated with a poor patient prognosis. However, therapeutic strategies to target these cells are difficult to implement due to the life-threatening risk of neutropenia. In a genetically engineered mouse model of lung adenocarcinoma, tumor-associated neutrophils (TAN) demonstrate tumor-supportive capacities and have a prolonged lifespan compared to circulating neutrophils. Here, we show that tumor cell-derived GM-CSF triggers the expression of the anti-apoptotic Bcl-xL protein and enhances neutrophil survival through JAK/STAT signaling. Targeting Bcl-xL activity with a specific BH3 mimetic, A-1331852, blocked the induced neutrophil survival without impacting their normal lifespan. Specifically, oral administration with A-1331852 decreased TAN survival and abundance, and reduced tumor growth without causing neutropenia. We also show that G-CSF, a drug used to combat neutropenia in patients receiving chemotherapy, increased the proportion of young TANs and augmented the anti-tumor effect resulting from Bcl-xL blockade. Finally, our human tumor data indicate the same role for Bcl-xL on pro-tumoral neutrophil survival. These results altogether provide preclinical evidence for safe neutrophil targeting based on their aberrant intra-tumor longevity.

Suggested role for neutrophil extracellular trap formation in Ewing sarcoma immune microenvironment.

Ewing sarcoma (EWS) is a highly aggressive cancer with a survival rate of 70%-80% for patients with localized disease and under 30% for those with metastatic disease. Tumor-infiltrating neutrophils (TIN) can generate extracellular net-like DNA structures known as neutrophil extracellular traps (NETs). However, little is known about the presence and prognostic significance of tumor-infiltrating NETs in EWS. Herein, we investigated 46 patients diagnosed with EWS and treated in the Tel Aviv Medical Center between 2010 and 2021. TINs and NETs were identified in diagnostic biopsies of EWS by immunofluorescence. In addition, NETs were investigated in neutrophils isolated from peripheral blood samples of EWS patients at diagnosis and following neoadjuvant chemotherapy. The relationships between the presence of TINs and NETs, pathological and clinical features, and outcomes were analyzed. Our results demonstrate that TIN and NETs at diagnosis were higher in EWS patients with metastatic disease compared with those with local disease. High NET formation at diagnosis predicted poor response to neoadjuvant chemotherapy, relapse, and death from disease (p < 0.05). NET formation in peripheral blood samples at diagnosis was significantly elevated among patients with EWS compared with pediatric controls and decreased significantly following neoadjuvant chemotherapy. In conclusion, NET formation seems to have a role in the EWS immune microenvironment. Their presence can refine risk stratification, predict chemotherapy resistance and survival, and serve as a therapeutic target in patients with EWS.

Neutrophils resist ferroptosis and promote breast cancer metastasis through aconitate decarboxylase 1

Metastasis causes breast cancer-related mortality. Tumor-infiltrating neutrophils (TINs) inflict immunosuppression and promote metastasis. Therapeutic debilitation of TINs may enhance immunotherapy, yet it remains a challenge to identify therapeutic targets highly expressed and functionally essential in TINs but under-expressed in extra-tumoral neutrophils. Here, using single-cell RNA sequencing to compare TINs and circulating neutrophils in murine mammary tumor models, we identified aconitate decarboxylase 1 (Acod1) as the most upregulated metabolic enzyme in mouse TINs and validated high Acod1 expression in human TINs. Activated through the GM-CSF-JAK/STAT5-C/EBPβ pathway, Acod1 produces itaconate, which mediates Nrf2-dependent defense against ferroptosis and upholds the persistence of TINs. Acod1 ablation abates TIN infiltration, constrains metastasis (but not primary tumors), bolsters antitumor Tcell immunity, and boosts the efficacy of immune checkpoint blockade. Our findings reveal how TINs escape from ferroptosis through the Acod1-dependent immunometabolism switch and establish Acod1 as a target to offset immunosuppression and improve immunotherapy against metastasis.