Tumor-associated Neutrophil Research Articles

ACSL6-activated IL-18R1-NF-κB promotes IL-18-mediated tumor immune evasion and tumor progression.

Aberrant activation of IL-18 signaling regulates tumor immune evasion and progression. However, the underlying mechanism remains unclear. Here, we report that long-chain acyl-CoA synthase 6 (ACSL6) is highly expressed in liver cancer and correlated with poor prognosis. ACSL6 promotes tumor growth, metastasis, and immune evasion mediated by IL-18, independent of its metabolic enzyme activity. Mechanistically, upon IL-18 stimulation, ACSL6 is phosphorylated by ERK2 at S674 and recruits IL-18RAP to interact with IL-18R1, thereby reinforcing the IL-18R1-IL-18RAP heterodimer and triggering NF-κB-dependent gene expression to facilitate tumor development. Furthermore, the up-regulation of CXCL1 and CXCL5 by ACSL6 promotes tumor-associated neutrophil and tumor-associated macrophage recruitment, thereby inhibiting cytotoxic CD8+ T cell infiltration. Ablation or S674A mutation of ACSL6 potentiated anti-PD-1 therapeutic efficacy by increasing the effector activity of intertumoral CD8+ T cells. We revealed that ACSL6 is a potential adaptor that activates IL-18-NF-κB axis-mediated tumor immune evasion and provides valuable insights for developing effective immunotherapy strategies for cancer.

A novel tumor-associated neutrophil gene signature for predicting prognosis, tumor immune microenvironment, and therapeutic response in breast cancer

Tumor-associated neutrophils (TANs) can promote tumor progression. This study aimed to investigate the molecular signature that predict the prognosis and immune response of breast cancer (BRCA) based on TAN-related gene (TANRG) expression data. The RNA-seq data of BRCA were gathered from The Cancer Genome Atlas (TCGA) and gene expression omnibus (GEO) datasets. Univariate Cox regression analysis and the least absolute shrinkage and selection operator for selecting prognostic genes. A neo-TAN-related risk signature was constructed by multivariate Cox regression analysis. Time-dependent receiver operating characteristic (ROC) curve analyses and Kaplan–Meier analyses were performed to validate the signature in GEO cohorts and the triple-negative breast cancer (TNBC) subtype. We constructed an independent prognostic factor model with 11 TANRGs. The areas under the ROC curve (AUCs) of the TCGA training cohorts for 3-, 5-, and 7-year overall survival were 0.72, 0.73, and 0.73, respectively. The AUCs of the GEO test cohorts for 3-, 5-, and 7-year overall survival were 0.83, 0.89, and 0.94 (GSE25066) and 0.67, 0.69, and 0.73 (GSE58812), respectively. The proportion of immune subtypes differed among the different risk groups. The IC50 values differed significantly between risk groups and can be used as a guide for systemic therapy. The prognostic model developed by TANRGs has excellent predictive performance in BRCA patients. In addition, this feature is closely related to the prediction of survival, immune activity and treatment response in BRCA patients.

CRKL dictates anti-PD-1 resistance by mediating tumor-associated neutrophil infiltration in hepatocellular carcinoma

The effectiveness of immune checkpoint inhibitor (ICI) therapy for hepatocellular carcinoma (HCC) is limited by treatment resistance. However, the mechanisms underlying immunotherapy resistance remain elusive. We aimed to identify the role of CT10 regulator of kinase-like (CRKL) in resistance to anti-PD-1 therapy in HCC. Gene expression in HCC specimens from 10 patients receiving anti-PD-1 therapy was identified by RNA-sequencing. A total of 404 HCC samples from tissue microarrays were analyzed by immunohistochemistry. Transgenic mice (Alb-Cre/Trp53fl/fl) received hydrodynamic tail vein injections of a CRKL-overexpressing vector. Mass cytometry by time of flight was used to profile the proportion and status of different immune cell lineages in the mouse tumor tissues. CRKL was identified as a candidate anti-PD-1-resistance gene using a pooled genetic screen. CRKL overexpression nullifies anti-PD-1 treatment efficacy by mobilizing tumor-associated neutrophils (TANs), which block the infiltration and function of CD8+ T cells. PD-L1+ TANs were found to be an essential subset of TANs that were regulated by CRKL expression and display an immunosuppressive phenotype. Mechanistically, CRKL inhibits APC (adenomatous polyposis coli)-mediated proteasomal degradation of β-catenin by competitively decreasing Axin1 binding, and thus promotes VEGFα and CXCL1 expression. Using human HCC samples, we verified the positive correlations of CRKL/β-catenin/VEGFα and CXCL1. Targeting CRKL using CRISPR-Cas9 gene editing (CRKL knockout) or its downstream regulators effectively restored the efficacy of anti-PD-1 therapy in an orthotopic mouse model and a patient-derived organotypic tumor spheroid model. Activation of the CRKL/β-catenin/VEGFα and CXCL1 axis is a critical obstacle to successful anti-PD-1 therapy. Therefore, CRKL inhibitors combined with anti-PD-1 could be useful for the treatment of HCC. Here, we found that CRKL was overexpressed in anti-PD-1-resistant hepatocellular carcinoma (HCC) and that CRKL upregulation promotes anti-PD-1 resistance in HCC. We identified that upregulation of the CRKL/β-catenin/VEGFα and CXCL1 axis contributes to anti-PD-1 tolerance by promoting infiltration of tumor-associated neutrophils. These findings support the strategy of bevacizumab-based immune checkpoint inhibitor combination therapy, and CRKL inhibitors combined with anti-PD-1 therapy may be developed for the treatment of HCC.

Knowledge landscape of tumor-associated neutrophil: a bibliometric and visual analysis from 2000-2024.

Neutrophils have long been consistently adjudged to hold a dominant position in acute inflammation, which once led people to undervalue their role in chronic malignancy. It is now acknowledged that neutrophils also infiltrate into the tumor microenvironment in substantial quantities and form a highly abundant immune population within the tumor, known as tumor-associated neutrophils (TANs). There has been a surge of interest in researching the eminent heterogeneity and plasticity of TANs in recent years, and scholars increasingly cotton on to the multifaceted functions of TANs so that strenuous endeavors have been devoted to enunciating their potential as therapeutic targets. Yet it remains much left to translate TAN-targeted immunotherapies into clinical practice. Therefore, there is great significance to comprehensively appraise the research status, focal point, and evolution trend of TAN by using bibliometric analysis. Publications related to TAN research from 2000 to 2024 are extracted from the Web of Science Core Collection. Bibliometric analysis and visualization were performed by tools encompassing Microsoft Excel, VOSviewer, CiteSpace, R-bibliometrix, and so on. The bibliometric analysis included a total of 788 publications authored by 5291 scholars affiliated with 1000 institutions across 58 countries/regions, with relevant articles published in 324 journals. Despite China's maximum quantity of publications and top 10 institutions, the United States is the leading country with the most high-quality publications and is also the global cooperation center. FRONTIERS IN IMMUNOLOGY published the most papers, whereas CANCER RESEARCH is the highest co-cited journal. Israeli professor Fridlender, Zvi G. is the founder, pioneer, and cultivator with the highest citation counts and H-index in the TAN area. Our analysis prefigures the future trajectories: TAN heterogeneity, neutrophil extracellular trap, the crosstalk between TANs and immunocytes, and immunotherapy will likely be the focus of future research. A comprehensive bibliometric and visual analysis is first performed to map the current landscape and intellectual structure of TAN, which proffers fresh perspectives for further research. The accurate identification of distinct TAN subpopulations and the precise targeting of key pro-tumor/anti-tumor subpopulations hold immense potential to develop into a TAN-targeted immunotherapy.

HPV-Negative and HPV-Positive Oral Cancer Cells Stimulate the Polarization of Neutrophils towards Different Functional Phenotypes In Vitro.

High-risk human papillomavirus (HPV) infection is one of the leading causes of oropharyngeal squamous cell carcinoma (OPSCC), while the correlation between HPV and oral squamous cell carcinoma (OSCC) remains controversial. The inflammatory infiltrate involved in these epithelial neoplasms differs based on their association with HPV. HPV- tumors show higher tumor-associated neutrophil (TAN) infiltration. It is believed that TANs can play a dual role in cancer by exerting either anti-tumorigenic or pro-tumorigenic effects. However, the impact of HPV status on neutrophil polarization remains unknown. Therefore, this study aimed to investigate the effect of OSCC cells, both HPV- and HPV16+, on the functional phenotype of neutrophils. Peripheral blood neutrophils were stimulated with supernatants from OSCC cell lines and non-tumorigenic HaCaT keratinocytes transduced with HPV16 E6/E7 oncogenes. Subsequently, cytokine production, cell viability, metabolism, expression of degranulation markers, and PD-L1 expression were evaluated. Our findings demonstrate that in contrast to UPCI:SCC154 (HPV+ OSCC) cells, the SCC-9 (HPV- OSCC) cell line induced a highly activated functional state in neutrophils, which is potentially associated with a pro-tumorigenic effect. The HaCaT 16-E7 supernatant only stimulated the activation of some neutrophil functions. Understanding the complex interplay between neutrophils and their microenvironment has the potential to identify TANs as viable therapeutic targets.

CellCharter reveals spatial cell niches associated with tissue remodeling and cell plasticity.

Tissues are organized in cellular niches, the composition and interactions of which can be investigated using spatial omics technologies. However, systematic analyses of tissue composition are challenged by the scale and diversity of the data. Here we present CellCharter, an algorithmic framework to identify, characterize, and compare cellular niches in spatially resolved datasets. CellCharter outperformed existing approaches and effectively identified cellular niches across datasets generated using different technologies, and comprising hundreds of samples and millions of cells. In multiple human lung cancer cohorts, CellCharter uncovered a cellular niche composed of tumor-associated neutrophil and cancer cells expressing markers of hypoxia and cell migration. This cancer cell state was spatially segregated from more proliferative tumor cell clusters and was associated with tumor-associated neutrophil infiltration and poor prognosis in independent patient cohorts. Overall, CellCharter enables systematic analyses across data types and technologies to decode the link between spatial tissue architectures and cell plasticity.

Yi Qi Chu Tan Formula (YQCTF) inhibited the progress of lung cancer via regulating tumor-associated neutrophil: An integrated study of network pharmacology, proteomics and pharmacodynamics

Ethnopharmacological relevanceYi Qi Chu Tan Formula (YQCTF), a prescription consisting of eight traditional Chinese medicine for treating lung cancer, has been clinically proven to be effective in improving the life quality and prolonging the survival time of non-small cell lung cancer (NSCLC) patients. Aim of the studyThis study aimed to evaluate the therapeutic efficacy of YQCTF on NSCLC mice model and further explore its therapeutic targets by using network pharmacology, proteomics and pharmacodynamic methodologies. Materials and methodsThe network pharmacology analysis was firstly conducted to screen out the potential active ingredients and therapeutic targets of YQCTF against NSCLC. Three kinds of extracts, i.e. the water extract (WE), water extraction-alcohol precipitation (WEAP) and alcohol extract (AE) of YQCTF were prepared, which chemical compositions were subsequently analyzed by using ultra-high performance liquid chromatography-mass spectrometry/mass spectrometry (UPLC-MS/MS), and which anti-neoplastic efficacy was examined on NSCLC mice model. Mice tumor tissues were collected for proteomics analysis, and the immunomodulatory effects of YQCTF extracts on the tumor microenvironment (TME) were further validated by using flow cytometry, immunofluorescence, ELISA and Western blot. ResultsNetwork pharmacology identified 60 conjunct genes and ample cancer-related signaling pathways as potential therapeutic targets of YQCTF. Protein-protein interaction (PPI), Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that YQCTF might negatively regulate cancer-related inflammation. UPLC-MS/MS analysis showed that the main components of YQCTF include at least ginsenosides, solasodine, solamargine, solasonine, peimisine, peiminine, peimine and sipeimine-3β-D-glucosihde. All kinds of YQCTF extracts significantly inhibited the growth of lung cancer allograft and regulated the ratio of immune cells in tumor tissues, i.e. upregulated the fractions of T cells, promoted the maturation of dendritic cells (DCs), increased the M1/M2 ratio of tumor-related macrophages, but reduced the number of Tregs and immunosuppressive neutrophils. Proteomics identified neutrophils to be the most prominently enriched target linked to NETs formation in mice tumor tissue, which is verified by the downregulation of neutrophil recruiting factors involving IL-6, HIF-1α and IL-8, as well as the decreases of NETs-related biomarkers including H3cit, MPO, CD18, MMP9 and ICAM-1 in immunofluorescence, ELISA and Western blot analysis. ConclusionYQCTF inhibited the progress of mice NSCLC allograft, suppressed the pro-tumorigenic tumor-associated neutrophils and improved the tumor immune microenvironment (TIME).

Simultaneous expression of androgen receptor and tumor-associated neutrophil in breast cancer subtypes.

e12539 Background: Considering the heterogeneity of breast cancer phenotypes and different response rates to treatment, it is essential to determine prognostic markers that could confer therapeutic role as well. In this study, androgen receptor (AR) and tumor-associated neutrophil (TAN) in female breast cancer were investigated and their coherency with different subtypes of breastcancer and pathology findings were assessed. Methods: This cross-sectional study was conducted on patients with Stage I-IV breast cancer who were referred to Borna Pathology Center in Zanjan, Iran during 2018-2021. The tumoral paraffine blocks were evaluated for AR (by the IHC method) and TAN (by morphology and the IHC staining for myeloperoxidase). Relationships in different subtypes and stages were assessed.Using univariate logistic regression between 5 subtypes (defended by ER, PR, HER2, and Ki67), stage, age, metastasis (M), and grade with AR and TAN were studied separately and simultaneously. Results: We collected data from 83 women with Stage I-IV breast cancer with mean ± SD age of diagnosis 49.3±11.2 years. Percentage of AR+, TAN+, and AR+&TAN+ were 71.1, 43.4, and26.5, respectively. The frequency of AR+ and AR-&TAN- was high in hormone-positive subtypes, while TAN+ was frequent in the triple-negative group and AR+&TAN+ in Luminal A and B groups. The relation between age, M, and Stage with AR, TAN, and AR & TAN was not significant. AR was significantly related to subtype and grade. Conclusions: AR+ probability was significantly lower in Triple Negative subtype and poorly differentiated grade. Based on the heterogeneity of the presentation of these markers, survival evaluation can be important in risk assessment of them. Relationship between AR, TAN, and AR&TAN with study variables. [Table: see text]

Relation assessment between TAN and AR with the survival of breast cancer patients.

e12542 Background: Considering the importance of the relationship between biomarkers and survival in breast cancer patients, finding this relationship can play a key role in understanding disease behavior, determining prognosis and even identifying targeted treatments. This study was conducted with the aim of investigating the relationship between TAN (Tumor associated neutrophil) and AR (Androgen receptor) with survival in patients with breast cancer. Methods: In this cohort study, 83 female patients with stage I- IV breast cancer were included. we investigated the relationship between TAN and AR with the three-year survival time. Patients were diagnosed between April 2018 and March 2021. Also, the latest follow-up was performed in December 2022. The Cox-hazard model was used to determine the factors influencing the survival time of patients. Results: Out of 83, follow-up was conducted for 81 women. The mean±SD age of diagnosis was 49.5±11.1 years. Sixteen (19.75%) of them died and the mean (95% C.I) for follow-up time was 32.78 (30.97-34.58) months. The one, two, and three survival rates were 0.95, 0.89, and 0.80, respectively. The mean survival time (month) for AR-TAN-, AR+TAN-, AR+TAN+, and AR-TAN+ was 35.75, 33.50, 32.36, and 28.58 respectively. The 3-year survival in the AR-TAN-, AR+TAN-, AR+TAN+ and AR-TAN+ groups was 0.88, 0.88, 0.79, and 0.50, respectively. Conclusions: AR positivity and TAN negativity are in favor of better three-year overall survival regardless of subtypes, however the differences were not statistically significant. The Survival status was significantly lower in the AR-TAN+ group compared to AR + TAN- group. These results are a promising way to the future studies. [Table: see text]

Apatinib remodels the immunosuppressive tumor ecosystem of gastric cancer enhancing anti-PD-1 immunotherapy

Apatinib has been shown to clinically enhance anti-PD-1 immunotherapy for advanced gastric cancer (GC). However, the complexity of GC immunosuppression remains a challenge for precision immunotherapy. Here, we profile the transcriptomes of 34,182 single cells from GC patient-derived xenografts of humanized mouse models treated with vehicle, nivolumab, or nivolumab plus apatinib. Notably, excessive expression of CXCL5 in the CellCycle malignant epithelium, induced by anti-PD-1 immunotherapy and blocked by combined apatinib treatment, is found to be a key driver of tumor-associated neutrophil (TAN) recruitment in the tumor microenvironment through the CXCL5/CXCR2 axis. We further show that the protumor TAN signature is associated with anti-PD-1 immunotherapy-related progressive disease and poor cancer prognosis. Molecular and functional analyses in cell-derived xenograft models confirm the positive invivo therapeutic effect of targeting the CXCL5/CXCR2 axis during anti-PD-1 immunotherapy. Altogether, our study elucidates the GC immunosuppressive landscape in anti-PD-1 immunotherapy and highlights potential targets for overcoming checkpoint immunotherapy resistance.

Abstract 5956: Tumor-infiltrating neutrophil progenitors impair homologous DNA repair and promote sensitivity to PARP-inhibition

Abstract Tumor evolution is one of the major mechanisms responsible for acquiring therapy-resistant and more aggressive cancer clones. Whether the tumor microenvironment through immune-mediated mechanisms might promote the development of more aggressive cancer types is crucial for the identification of additional therapeutical opportunities. Here, we identified a novel subset of tumor-infiltrating neutrophils, defined as tumor-associated neutrophil progenitors (NePs). These NePs are enriched in highly proliferative hormone-dependent breast cancers and impair DNA repair capacity. Mechanistically, succinate secreted by tumor-associated NePs inhibits homologous recombination, promoting error-prone DNA repair through non-homologous end-joining regulated by PARP-1. Consequently, breast cancer cells acquire genomic instability, resulting in tumor progression and resistance to endocrine therapies. Selective inhibition of these pathways induces increased tumor cell kill in vitro and in vivo. Intra-tumor NePs score correlates with copy number alterations in highly proliferative hormone-dependent tumors from breast cancer patients. Treatment with PARP-1 inhibitors counteract the pro-tumorigenic effect of these neutrophils and reverses endocrine resistance. Citation Format: Arianna Calcinotto. Tumor-infiltrating neutrophil progenitors impair homologous DNA repair and promote sensitivity to PARP-inhibition. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5956.

ETV4 Mediated Tumor‐Associated Neutrophil Infiltration Facilitates Lymphangiogenesis and Lymphatic Metastasis of Bladder Cancer (Adv. Sci. 11/2023)

Tumor-Associated Neutrophil In article number 2205613, Xu Chen, Tianxin Lin, and co-workers reveal a novel mechanism of tumor-associated neutrophil (TANs) in lymph node (LN)-metastatic bladder cancer. ETS variant transcription factor (ETV4) enhances bladder cancer (BCa) cells-derived chemokine C-X-C motif ligand (CXCL)1/8 to recruit TANs, leading to the increase of vascular endothelial growth factor A (VEGFA) and matrix metalloproteinase-9 (MMP9), and then facilitating lymphangiogenesis and LN metastasis of bladder cancer.

Tumor Associated Neutrophil Polarization and its Potential Effects in Cancer Therapy

Neutrophils are the first line of defense against pathogens in the immune system, and also has important implications for the tumor micro-environment. The effect of neutrophils on cancer is a double-edged sword in that they can both promote or inhibit tumor growth. When in different tumor environments or exposed to different stimulation, neutrophils would polarize toward anti-tumorigenic N1 or pro-tumorigenic N2 phenotype. The exact mechanism to polarize N1 and N2 are not yet well understood, but it is currently known that IFN-β or stimulation of the pathogen-associated molecular patterns (PAMPs) pathway can cause N1 polarization. On the contrary, in the presence of TGF-β or IL-10, N2 polarization occurs. When N1 population is dominant in the tumor micro-environment, they will inhibit tumor growth and enhance the effect of immunotherapy. But vice versa, dominance of pro-tumorigenic N2 phenotype within the tumor would enhance inflammation leading to more aggressive tumor growth and also hinders the effect of immunotherapy. Therefore, it is very important to understand the subtype of tumor associated neutrophils in the tumor when determining appropriate cancer therapy.

Prognostic value of tumor-associated N1/N2 neutrophil plasticity in patients following radical resection of pancreas ductal adenocarcinoma

BackgroundAs an integral part of the tumor microenvironment (TME), tumor-associated neutrophils play a crucial role in tumor development. The objective of this study was to investigate the plasticity of tumor-associated...

Single-cell RNA sequencing analysis revealed cellular and molecular immune profiles in lung squamous cell carcinoma

Although breakthroughs have been made in the treatment of non-small cell lung cancer, there are only a few choices for advanced-stage or recurrent lung squamous cell carcinoma (LUSC) patients. In our study, we identified 7 major cell types in thedepicted the immunolandscape of LUSC microenvironment using single-cell RNA sequencing. We found that an immunosuppressive receptor, T cell immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domain (TIGIT), was highly expressed by regulatory T cells (Tregs) and exhausted CD8+T cells, suggesting that upregulation of TIGIT might promote an immunosuppressive microenvironment and inhibit the cytotoxic ability of CD8+T cells. We also identified tumor-associated neutrophil (TAN), characterized by CXCR2, CSF3R and CXCL8, in the tumor region, and TANs upregulated the expression of interleukin 1 receptor antagonist (IL1RN) which suggested that TAN might exert an immunosuppressive role via expressing IL1RN. Furthermore, the number of SPP1+ macrophages(SPP1+M) significantly increased in tumor microenvirnment, which was correlated with the poor survival of patients. Additionally, regulatory networks based on SPP1+M revealed that the disparities of several ligand-receptor pairs existed between tumor and normal tissues. Among these pairs, SPP1-CD44 showed the most interactions between SPP1+M and other cell types. Our results provided deep insight into the immune landscape of LUSC and an essential resource for drug discovery in the future.

High PANX1 Expression Leads to Neutrophil Recruitment and the Formation of a High Adenosine Immunosuppressive Tumor Microenvironment in Basal-like Breast Cancer.

Simple SummaryA high adenosine level is an important characteristic of the tumor microenvironment (TME) in breast cancer. Pannexin 1 (PANX1) can release intracellular ATP to the extracellular space and elevate extracellular ATP (exATP) levels under physiological conditions. PANX1 has been found to be a poor prognostic factor in breast cancer, however, the role of PANX1 in breast cancer remains unknown. In this study, we performed RNA sequencing, bioinformatics analysis, surgical specimen histological validation, and exATP/extracellular adenosine (exADO) assays to reveal the role of PANX1 in regulating the immune microenvironment of basal-like breast cancer. The results revealed that PANX1 acted as a poor prognostic factor for breast cancer and had high expression in basal-like breast cancer. PANX1 expression was positively correlated with exATP and exADO levels in basal-like breast cancer TME. PANX1 expression was also positively correlated with tumor-associated neutrophil (TAN) infiltration in breast cancer TME, and TANs highly expressed CD39/CD73, which synergistically build a high exADO immunosuppressive TME and promote tumor progression. This study suggests that high PANX1 expression is associated with high TAN infiltration and adenosine production to induce local immunosuppression in basal-like breast cancer TME.Background: A high adenosine level is an important characteristic of the tumor microenvironment (TME) in breast cancer. Pannexin 1 (PANX1) can release intracellular ATP to the extracellular space and elevate extracellular ATP (exATP) levels under physiological conditions. Methods: We performed public database bioinformatics analysis, surgical specimen histological validation, RNA sequencing, and exATP/extracellular adenosine (exADO) assays to reveal the role of PANX1 in regulating the immune microenvironment of basal-like breast cancer. Results: Our results revealed that PANX1 acted as a poor prognostic factor for breast cancer and had high expression in basal-like breast cancer. PANX1 expression was positively correlated with exATP and exADO levels in basal-like breast cancer TME. PANX1 expression was also positively correlated with tumor-associated neutrophil (TAN) infiltration in breast cancer TME and TANs highly expressed ENTPD1 (CD39)/NT5E (CD73). Conclusions: This study suggests that high PANX1 expression is associated with high TAN infiltration and adenosine production to induce local immunosuppression in basal-like breast cancer TME.

Abstract 5266: Prognostic role of molecular subclassification based on mutational profiling and tumor-associated neutrophil status in stage III colon cancer

Abstract The traditional prognostic prediction of colon cancer (CC) is based on the tumor-node-metastasis (TNM) system. This, however, is not adequate for clinical decision making due to the existence of inter-patient variations and heterogeneity of underlying molecular pathways. We aim to investigate the prognostic value of molecular subclassification utilizing i) tumor-associated neutrophil (TAN) status by immunohistochemical (IHC) staining; ii) microsatellite instability (MSI) and elevated microsatellite alteration at selected tetranucleotide repeats (EMAST) status by PCR-based microsatellite typing; and iii) next-generation sequencing (NGS) mutational profiling by deep sequencing of a 77-gene oncology panel. In the discovery phase, 130 pairs of archived normal and tumor formalin-fixed paraffin-embedded (FFPE) specimens of stage III CC patients were retrospectively recruited from Queen Mary Hospital. CC patients will be categorized into different molecular subclasses (MSCs) based on the status of TAN/MSI/EMAST/NGS mutational profiling molecular biomarkers. Kaplan-Meier analysis, log rank test, univariate and multivariate Cox proportional hazards regression analysis will be used for estimation of progression-free survival (PFS) and overall survival (OS) between the two groups of CC patients. We expect our data to provide useful information for improved stratification of stage III CC patients into high or low risk of recurrence and survival. Acknowledgement: The Health and Medical Research Fund (07181536) to JMYK. Citation Format: CY Chan, AWI Lo, KO Lam, HY Ng, CC Chen, ML Lung, JMY Ko. Prognostic role of molecular subclassification based on mutational profiling and tumor-associated neutrophil status in stage III colon cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5266.

Abstract 4213: Targeting the cMet inhibitors combined with anti-PD-1 therapy: Preclinical approach to turn the resistance into opportunities for overcoming tumor evasion

Abstract Liver cancer is the fourth most common cause of cancer-related mortality world-wide, and Hepatocellular carcinoma (HCC) is the most common form of liver cancer accounting for 90% of cases. Therapeutic strategies include surgical resection, radiotherapy, liver transplantation and pharmacological treatment with small-molecules multikinase inhibitors and/or immune-checkpoint blockade (ICB); however, outcomes remain poor and improvements to systemic therapy are desperately needed. Recently, Cabozantinib a kinase inhibitor with activity against VEGFR2, AXL and cMET, has been approved for second line use in advanced HCC; however, monotherapy achieves less than a 4-month extension of overall survival. Previously, our collaborators demonstrated that the efficacy of cMet inhibition is self-limited by induction of high PD-L1 expression. Given this data, the goal of our study was to investigate the efficacy of combining cMet inhibition with either capmatinib or cabozantinib with antibody blockade of the PD-1 immune checkpoint receptor for therapy of HCC. Using multiple syngeneic, immunocompetent models of HCC (Hepa1-6, HCA) in two different strains of mice (C57BL6, C3H), we investigated the combination potential of each of these cMet inhibitors with anti-PD-1 antibodies. Further, we profiled the changes to the tumor infiltrating myeloid and T cell repertoires which correlated to mono- as well as combination therapy responses in these animals using high content flow cytometry on a BD X-30. We found Hepa1-6 to be exquisitely anti-PD-1 sensitive with 100% of animals being cured by the combination. In the less immunotherapy-sensitive HCA-1 model, we also found that the cMet inhibitors cooperate with PD-1 blockade to promote rejection of HCA-1 HCC model. Again, we found that combination therapy could cure nearly all animals of pre-implanted HCA-1 tumors. Mechanistically, we found that concomitant cMet inhibition enhances the efficacy of PD-1 therapy increasing the number of tumor-infiltrating CD4 Teff cells while decreasing the CD4 Treg population. We also found that combined therapy improves CD8+ T cell infiltration, memory CD8 T cell fraction and NK cytotoxic potential within the tumor microenvironment. Within the HCA-1 myeloid compartment, we discovered a significant decrease in the tumor-associated neutrophil (N2-TAN) population (PMN-MDSC). Further, we found that combination therapy promotes pro-inflammatory M1 conversion of stromal macrophages. Collectively, these data provide a rationale for combining anti-PD-1 therapy with cMet inhibitors as a potential strategy to overcome resistance to immune checkpoint blockade therapy in HCC. Citation Format: Ricardo Alexandre de Azevedo, Hsieh Cheng-En, Priyamvada Jayaprakash, Krithikaa Rajkumar Bhanu, Anupallavi Srinivasamani, Brittany Morrow, Michelle Winkler, Shweta Mahendra Hedge, Arthur Liu, Michael A. Curran. Targeting the cMet inhibitors combined with anti-PD-1 therapy: Preclinical approach to turn the resistance into opportunities for overcoming tumor evasion [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 4213.

Clinicopathological Characteristics of Cervical Squamous Cell Carcinoma and Correlation with Tumor-Associated Neutrophil

BackgroundCervical carcinoma is one of the most common cancers in women, according to data from World Health Organization (WHO).Cervical carcinoma ranks the fourth most deadly type of cancer. Squamous cell carcinoma is the most common type of this cervicalcancer and it is generally caused by Human Papilloma Virus (HPV) types 16 and 18. Tumor-Associated Neutrophil (TAN) are foundin various types of cancer, the density of TAN can determine the prognostic type of tumor or cancer in each individual. This studywas aimed to identify correlation between clinicopathological characteristics of cervical squamous cell carcinoma (SCC) and TAN incervical SCC patients registered at RSUP Dr. Mohammad Hoesin Palembang.MethodsThis type of research is analytic with a cross-sectional method. Data collection in this study used a 37 slides archive which werediagnosed as cervical squamous cell carcinoma in Anatomic Pathology in RSUP Dr. Mohammad Hoesin Palembang for the periodof 2017-2019.ResultsFrom 37 cervical SCC patients, the clinicopathological characteristics of the majority of cervical SCC patients occurred in the agegroup of 51 year-old (51.4%), with histopathological type of non-keratinizing SCC (75,7%) and the most frequent differentiationdegree was moderately differentiated (75,7%). The TAN density was mostly found at low TAN densities (73%). There wassignificant correlation between age and TAN density (p=0.035) and there was no significant correlation between age and TANdensity (p=0.269).ConclusionIt was explained that the higher the degree of differentiation, the lower the density value, however the neutrophils in this studycannot be ascertained whether the neutrophils of anti-tumor (N1) or pro-tumor (N2) characteristic

Abstract P5-06-05: Palmitate induces a senescent-like phenotype in fibroblasts resulting in altered phenotypes in cells of the breast tumor microenvironment

Abstract Background: Obese breast cancer patients face a worse prognosis, including an increased risk of recurrence and mortality. While the causative mechanisms have yet to be fully uncovered, emerging evidence implicates palmitate, increased in the obese state, in development of cellular senescence, an inflammatory state associated with proliferation, metastasis, and tumor-associated neutrophil (TAN) polarization, among other measures of carcinogenesis. However, studies have yet to investigate the impact of palmitate on induction of senescence in breast fibroblasts, and no studies have assessed the effect of senescent fibroblasts on neutrophil polarization in the breast tumor microenvironment. This said, we hypothesize that palmitate alters breast cancer cell gene expression and neutrophil phenotype via induction of a senescent-like phenotype in fibroblasts. Methods: HCA2, IMR-90, and human mammary fibroblasts were exposed to palmitate or vehicle in media supplemented with 2% charcoal-stripped fetal bovine serum, after which the cells were measured through qPCR for expression of IL-1a, IL-6 and IL-8, some of the most prominent members of the senescence-associated secretory phenotype. Palmitate-exposed fibroblasts were also subjected to chromogenic staining for senescence-associated beta-galactosidase and immunoenzymatic BrdU analysis, two well-established measures of senescence. Next, in order to study the influence of these fibroblasts on other cells of the breast tumor microenvironment, we assessed their impact on polarization of DMSO-differentiated HL-60 neutrophils by using flow cytometry to measure neutrophil expression of CD54 and CD95, differentially expressed on N1 and N2 neutrophils. Finally, we employed PCR arrays to assess the impact of palmitate-exposed fibroblasts on the expression of 84 genes in MCF-7 and 231 breast cancer cells, measuring activation of pathways related to apoptosis, cell cycle, DNA damage, senescence, telomere maintenance, metabolism, angiogenesis, and the endothelial-to-mesenchymal transition. Results and Conclusions: Palmitate induced pro-inflammatory gene expression and SA-beta-gal activity and decreased BrdU incorporation in fibroblasts. Palmitate also exhibited non-cell-autonomous effects, as palmitate-exposed fibroblasts induced phenotypic changes in both neutrophils and breast cancer cells. These findings are among the first to implicate palmitate-induced fibroblast senescence in the stimulation of non-cell-autonomous changes in the breast tumor microenvironment and will ultimately inform our understanding of the mechanistic connection between the obesity-associated factor palmitate and breast tumorigenesis. Citation Format: Brittany Susanne Harlow, Albert Davalos, Bryan McClellan, Andrew Brenner, Christopher Jolly, Stefano Tiziani, Steve Hursting, Linda deGraffenried. Palmitate induces a senescent-like phenotype in fibroblasts resulting in altered phenotypes in cells of the breast tumor microenvironment [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-06-05.