scholarly journals Irradiation suppresses STAT3-mediated MCL1 expression to augment CD8+ T cells cytotoxicity against EGFR-positive lung cancer

2021 ◽  
Author(s):  
Chun-I Wang ◽  
Yi-Fang Chang ◽  
Zong-Lin Sie ◽  
Ai-Sheng Ho ◽  
Chun-Chia Cheng
Keyword(s):  
Gene Expression ◽  
Lung Cancer ◽  
T Cells ◽  
Tumor Cells ◽  
Immune Surveillance ◽  
A549 Cells ◽  
Irradiation Effect ◽  
Small Cell Lung ◽  
Western Blots ◽  
Stat3 Phosphorylation

Abstract Background Tumor cells progress to evade immunological attacks and prohibit activity of CD8+ T cells. Irradiation damages tumor cells and augments tumor immunotherapy in clinical application. However, the detail mechanism remains elusive. We aimed to uncover the mechanism of irradiation augmenting cytotoxic CD8+ T cells to suppress tumor progression in non-small-cell lung cancer (NSCLC). Methods EGFR-positive NSCLC cell lines were co-cultured with isolated PBMCs from healthy volunteers, cell viability and apoptosis were measured. RNAseq was used to screen the IFNγ-mediated gene expression in A549 cells. Irradiation was used to augment PBMCs-mediated anti-tumor effect and the irradiation effect to IFNγ-mediated gene expression was investigated using qPCR and Western blots. Results Co-culture of tumor cells stimulates increase of granzyme B and IFNγ in CD8+ T, but A549 exhibits resistance against CD8+ T cytotoxicity. Irradiation inhibits A549 proliferation and enhances apoptosis, augmenting PBMCs-mediated cytotoxicity against A549. IFNγ simultaneously increased phosphorylation on STAT1 and STAT3 in EGFR-positive lung cancer, resulting in overexpression of PD-L1. In RNAseq analysis, MCL1 was identified and increased by IFNγ-STAT3 axis in A549 cells, we found that irradiation specifically inhibits phosphorylation on STAT1 and STAT3 in IFNr-treated A549, resulting in reductions of PD-L1 and MCL1. Moreover, knockdowns of STAT3 and MCL1 increased PBMCs against irradiated A549 cells. Conclusion This study demonstrated that A549 expressed MCL1 against CD8+ T cell-mediated apoptosis. In addition, we found that irradiation suppressed STAT3 phosphorylation and IFNγ-mediated PD-L1 and MCL1 expression, revealing a potential mechanism of irradiation augmenting immune surveillance.

scholarly journals Irradiation Suppresses IFNγ-Mediated PD-L1 and MCL1 Expression in EGFR-Positive Lung Cancer to Augment CD8+ T Cells Cytotoxicity

Cells ◽  
2021 ◽  
Vol 10 (10) ◽  
pp. 2515
Author(s):  
Chun-I. Wang ◽  
Yi-Fang Chang ◽  
Zong-Lin Sie ◽  
Ai-Sheng Ho ◽  
Jung-Shan Chang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Lung Cancer ◽  
T Cells ◽  
T Cell ◽  
Tumor Cells ◽  
Cd8 T Cells ◽  
Cd8 T Cell ◽  
Irradiation Effect ◽  
Western Blots ◽  
Stat3 Phosphorylation

Tumor cells express immune checkpoints to exhaust CD8+ T cells. Irradiation damages tumor cells and augments tumor immunotherapy in clinical applications. However, the radiotherapy-mediated molecular mechanism affecting CD8+ T cell activity remains elusive. We aimed to uncover the mechanism of radiotherapy augmenting cytotoxic CD8+ T cells in non-small-cell lung cancer (NSCLC). EGFR-positive NSCLC cell lines were co-cultured with CD8+ T cells from healthy volunteers. Tumor cell viability and apoptosis were consequently measured. IFNγ was identified secreted by CD8+ T cells and PBMCs. Therefore, RNAseq was used to screen the IFNγ-mediated gene expression in A549 cells. The irradiation effect to IFNγ-mediated gene expression was investigated using qPCR and western blots. We found that the co-culture of tumor cells stimulated the increase of granzyme B and IFNγ in CD8+ T, but A549 exhibited resistance against CD8+ T cytotoxicity compared to HCC827. Irradiation inhibited A549 proliferation and enhanced apoptosis, augmenting PBMCs-mediated cytotoxicity against A549. We found that IFNγ simultaneously increased phosphorylation on STAT1 and STAT3 in EGFR-positive lung cancer, resulting in overexpression of PD-L1 (p < 0.05). In RNAseq analysis, MCL1 was identified and increased by the IFNγ-STAT3 axis (p < 0.05). We demonstrated that irradiation specifically inhibited phosphorylation on STAT1 and STAT3 in IFNγ-treated A549, resulting in reductions of PD-L1 and MCL1 (both p < 0.05). Moreover, knockdowns of STAT3 and MCL1 increased the PBMCs-mediated anti-A549 effect. This study demonstrated that A549 expressed MCL1 to resist CD8+ T cell-mediated tumor apoptosis. In addition, we found that irradiation suppressed IFNγ-mediated STAT3 phosphorylation and PD-L1 and MCL1 expression, revealing a potential mechanism of radiotherapy augmenting immune surveillance.

scholarly journals CD8+ PD-1+ T-cells and PD-L1+ circulating tumor cells in chemotherapy-naïve non-small cell lung cancer: towards their clinical relevance?

2019 ◽  
Vol 11 ◽  
pp. 175883591985319 ◽  
Author(s):  
Athanasios Kotsakis ◽  
Galatea Kallergi ◽  
Despoina Aggouraki ◽  
Zaharoula Lyristi ◽  
Filippos Koinis ◽  
...  
Keyword(s):  
Lung Cancer ◽  
T Cells ◽  
Small Cell Lung Cancer ◽  
Tumor Cells ◽  
Circulating Tumor Cells ◽  
Cell Lung Cancer ◽  
Clinical Relevance ◽  
Small Cell ◽  
Response To Treatment ◽  
Small Cell Lung

Background: Since tumor cells may escape from immune surveillance through the programmed cell death 1 (PD-1)/programmed death ligand (PD-L)1 axis, this study was designed in order to evaluate whether there is a correlation between the levels of PD-1+ and PD-L1+-expressing immune cells (ICs) and circulating tumor cells (CTCs) in patients with non-small cell lung cancer (NSCLC). Patients and methods: Peripheral blood was obtained from 37 chemotherapy-naïve patients with metastatic NSCLC before treatment. PD-1 and PD-L1 expression was evaluated (1) on ICs with anti-tumor function (CD4+ and CD8+ T-cells, B-cells, monocytes/dendritic cells) using flow cytometry, (2) on CTCs by immunofluorescence and (3) on cells from tumor tissues by immunohistochemistry. The levels of PD-1+ and PD-L1+-expressing ICs were correlated with progression-free survival (PFS). Results: The presence of PD-1+ CD8+ cells, with reduced interferon (IFN)-γ expression, but not other ICs, were positively correlated with PD-L1+ CTCs ( p < 0.04). Increased percentages of PD-1+ CD8+ T-cells, were associated with a worse response to treatment ( p = 0.032) and shorter PFS ( p = 0.023) which, in multivariate analysis, was revealed as an independent predictor for decreased PFS [hazard ratio (HR): 4.1, p = 0.0007]. Conclusion: The results of the current study, for first time, provide evidence for a possible interaction between ICs and CTCs in NSCLC patients via the PD-1/PD-L1 axis and strongly support that the levels of PD-1+ CD8+ in these patients may be of clinical relevance.

The individuality of tumor-stroma interaction in non-small cell lung cancer: Insights from functional and molecular analyses

2009 ◽  
Vol 27 (15_suppl) ◽  
pp. 11115-11115
Author(s):  
S. Gottschling ◽  
R. Kuner ◽  
M. Granzow ◽  
E. Chang Xu ◽  
T. Muley ◽  
...  
Keyword(s):  
Gene Expression ◽  
Lung Cancer ◽  
Small Cell Lung Cancer ◽  
Tumor Cells ◽  
Cell Lung Cancer ◽  
Stromal Cells ◽  
Tumor Stroma ◽  
Small Cell ◽  
Normal Lung ◽  
Small Cell Lung

11115 Background: Tumor-stroma interaction plays a significant role for malignant growth. Results from prostate and breast cancer rodent models show cancerogenic properties of tumor-associated and genetically altered stromal cells (SC) when combined with initiated or normal epithelium (Olumi et al., Cancer Res 1999, Kuperwasser et al., PNAS 2004). However, data on the mechanisms and sequels of tumor-stroma interaction in lung cancer are scanty. Methods: Here, we analyzed the functional and molecular sequels of cross-talk between the non-small cell lung cancer (NSCLC) cell lines A549, H23, and H1703 and primary stromal cells (SC) derived from matched normal lung tissue and tumors of newly diagnosed NSCLC patients. Tumor cells were kept in a non-contact co-culture system with SC and analyzed for alterations in proliferation, colony formation, migration, adhesion, invasion, chemosensitivity and gene expression by Affymetrix HG U133 Plus 2.0 arrays. Results: Exposure to SC altered cellular functions and gene expression profiles related to tumor growth, metastasis and response to therapy. Each cell line showed individual alterations that were hierarchically governed by the (1) type of tumor cell, (2) the SC donor and his histology (3) and the local origin of the SC (normal lung tissue vs. tumor-associated). Conclusions: This in vitro model demonstrates an individual pattern of tumor-stroma interaction in NSCLC that is determined by both, the properties of the tumor cells and those of the stromal environment. Thus, biomarker programs in NSCLC should also consider the stromal compartment. No significant financial relationships to disclose.

scholarly journals Programmed cell death ligand-1-mediated enhancement of hexokinase 2 expression is inversely related to T-cell effector gene expression in non-small-cell lung cancer

2019 ◽  
Vol 38 (1) ◽  
Author(s):  
Sehui Kim ◽  
Ji-Young Jang ◽  
Jaemoon Koh ◽  
Dohee Kwon ◽  
Young A. Kim ◽  
...  
Keyword(s):  
Lung Cancer ◽  
T Cells ◽  
T Cell ◽  
Small Cell Lung Cancer ◽  
Tumor Cells ◽  
Cell Lung Cancer ◽  
Small Cell ◽  
Metabolic Reprogramming ◽  
Small Cell Lung ◽  
Cell Effector

Abstract Background We investigated the role of PD-L1 in the metabolic reprogramming of non-small cell lung cancer (NSCLC). Methods Changes in glycolysis-related molecules and glycolytic activity were evaluated in PD-L1low and PD-L1high NSCLC cells after transfection or knockdown of PD-L1, respectively. Jurkat T-cell activation was assessed after co-culture with NSCLC cells. The association between PD-L1 and immune response-related molecules or glycolysis were analyzed in patients with NSCLC and The Cancer Genome Atlas (TCGA). Results Transfecting PD-L1 in PD-L1low cells enhanced hexokinase-2 (HK2) expression, lactate production, and extracellular acidification rates, but minimally altered GLUT1 and PKM2 expression and oxygen consumption rates. By contrast, knocking-down PD-L1 in PD-L1high cells decreased HK2 expression and glycolysis by suppressing PI3K/Akt and Erk pathways. Interferon-γ (IFNγ) secretion and activation marker expression was decreased in stimulated Jurkat T-cells when co-cultured with HK2-overexpressing vector-transfected tumor cells rather than empty vector-transfected tumor cells. Immunohistochemistry revealed that PD-L1 expression was positively correlated with HK2 expression in NSCLC (p < 0.001). In TCGA, HK2 exhibited a positive linear association with CD274 (PD-L1) expression (p < 0.001) but an inverse correlation with the expression of CD4, CD8A, and T-cell effector function-related genes in the CD274high rather than CD274low group. Consistently, there were fewer CD8+ T-cells in PD-L1positive/HK2high tumors compared to PD-L1positive/HK2low tumors in squamous cell carcinoma. Conclusions PD-L1 enhances glycolysis in NSCLC by upregulating HK2, which might dampen anti-tumor immunity. PD-L1 may contribute to NSCLC oncogenesis by inducing metabolic reprogramming and immune checkpoint.

Correlation of circulating tumor enumeration and immune checkpoint marker expression on CD103+, CD4+ T cells in non-small cell lung cancer tissue.

2019 ◽  
Vol 37 (8_suppl) ◽  
pp. 109-109
Author(s):  
Xiaoyang WANG ◽  
Pin-I Chen ◽  
Maria Jaimes ◽  
Humin Gu ◽  
Keith Shults ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Flow Cytometry ◽  
T Cells ◽  
Tumor Cells ◽  
Immune Checkpoint ◽  
Clustering Analysis ◽  
Nsclc Patient ◽  
Small Cell ◽  
Small Cell Lung ◽  
Cancer Markers

109 Background: Non-small cell lung cancer (NSCLC) has a poor prognosis as most patients are at advanced stage when diagnosed. Targeted therapy and immunotherapy in recent years has significantly improved NSCLC patient outcome. In this study, we employed cell-by-cell immune and cancer marker profiling of the primary tumor cells to investigate possible signatures that might predict the presence or absence of circulating tumor cells (CTCs). Methods: We performed a comprehensive study on 10 NSCLC patient tissue samples with paired blood samples. The solid tissue biopsy samples were dissociated into single cells by non-enzymatic tissue homogenization. The single cell suspensions were stained with a total 25 immune, cancer markers and a DNA content dye and analyzed with advanced, high-parameter flow cytometry. CTCs were isolated and analyzed from the paired peripheral blood. Results: Out of the 26 unique cell markers stained, we investigated a total of 72 biomarkers for their correlation with CTC number. Strong correlations were observed between CTC number and the frequency of immune checkpoint marker expressing lymphocytes, especially with the immune checkpoint marker expressing CD103+CD4+ T lymphocytes. CTC number is also correlated with the frequency of PD-L1 expressing cancer cells and cancer cell DNA content. In contrast, CTC number inversely correlated to the frequency of CD44+E-cadherin- cancer cells. Unsupervised clustering analysis based on the biomarker analysis separated the CTC negative patients from the CTC positive patients. Conclusions: Profiling multiple immune and cancer markers on cancer samples with multi-parametric flow cytometry allowed us to obtain protein expression information at the single cell level. Clustering analysis of the proteomic data revealed a signature driven by checkpoint marker expression on CD103+CD4+ T cells that could potentially be predictive of CTCs.

Comprehensive Bioinformatics Analysis Identifies Tumor Microenvironment and Immune-related Genes in Small Cell Lung Cancer

2020 ◽  
Vol 23 (5) ◽  
pp. 381-391 ◽  
Author(s):  
Yongchun Song ◽  
Yanqin Sun ◽  
Tuanhe Sun ◽  
Ruixiang Tang
Keyword(s):  
Gene Expression ◽  
Lung Cancer ◽  
T Cells ◽  
Tumor Microenvironment ◽  
Immune Cells ◽  
Molecular Subtypes ◽  
Small Cell ◽  
Small Cell Lung ◽  
Immune Genes

Background: Tumor microenvironment (TME) cells play important roles in tumor progression. Accumulating evidence show that they can be exploited to predict the clinical outcomes and therapeutic responses of tumor. However, the role of immune genes of TME in small cell lung cancer (SCLC) is currently unknown. Objective: To determine the role of immune genes in SCLC. Methods: We downloaded the expression profile and clinical follow-up data of SCLC patients from Gene Expression Omnibus (GEO), and TME infiltration profile data of 158 patients using CIBERSORT. The correlation between TME phenotypes, genomic features, and clinicopathological features of SCLC was examined. A gene signature was constructed based on TME genes to further evaluate the relationship between molecular subtypes of SCLC with the prognosis and clinical features. Results: We identified a group of genes that are highly associated with TME. Several immune cells in TME cells were significantly correlated with SCLC prognosis (p<0.0001). These immune cells displayed diverse immune patterns. Three molecular subtypes of SCLC (TMEC1-3) were identified on the basis of enrichment of immune cell components, and these subtypes showed dissimilar prognosis profiles (p=0.03). The subtype with the best prognosis, TMEC3, was enriched with immune activation factors such as oncogene M0, oncogene M2, T cells follicular helper, and T cells CD8 (p<0.001). The TMEC1 subtype with the worst prognosis was enriched with T cells CD4 naive, B cells memory and Dendritic cells activated cells (p<0.001). Further analysis showed that the TME was significantly enriched with immune checkpoint genes, immune genes, and immune pathway genes (p<0.01). From the gene expression data, we identified four TME-related genes, GZMB, HAVCR2, PRF1 and TBX2, which were significantly associated with poor prognosis in both the training set and the validation set (p<0.05). These genes may serve as markers for monitoring tumor responses to immune checkpoint inhibitors. Conclusion: This study shows that TME features may serve as markers for evaluating response of SCLC cells to immunotherapy.

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