Low Infiltration Group Research Articles

Examining the influence of tumor-infiltrating macrophages on breast cancer outcomes and identifying relevant genes for diagnostic purposes

ObjectiveThe purpose of this research was to investigate how different types of immune cells impact the outlook of individuals with breast cancer, as well as identify the essential genes associated with immune cell subtype enrichment.MethodsThe Cancer Genome Atlas (TCGA) database and Gene Expression Omnibus (GEO) database were used to obtain global transcriptome sequencing data sets of breast tissue. The study utilized the CIBERSORT algorithm to determine the presence of 22 different types of immune cells in both breast cancer tissue and normal breast tissue.Immune cell infiltration content was utilized to conduct univariate COX analysis in order to identify risk factors linked to breast cancer prognosis.ResultsUnivariate COX analysis indicates that Macrophages M1 and B cells naive are beneficial factors for the outlook of individuals with breast cancer (P < 0.05), while Macrophages M2 and Monocytes are detrimental factors for the prognosis of breast cancer patients (P < 0.05). The high infiltration group of macrophage M2 had a poorer prognosis compared to the low infiltration group (P < 0.001); Conversely, the high infiltration group of macrophage M1 had a better prognosis than the low infiltration group (P = 0.002).ConclusionThe study provided an overview of immune cell infiltration in breast cancer tissues, identifying macrophage M1 and macrophage M2 as potential factors in breast cancer development and progression. Additionally, genes associated with macrophage phenotype were analyzed, offering insights into macrophage polarization mechanisms.

Genomics and tumor microenvironment of breast mucoepidermoid carcinoma based on whole-exome and RNA sequencing

Mammary mucoepidermoid carcinoma (MEC) is a rare entity. The molecular characteristics of breast MEC have not been fully investigated due to its rarity. We performed a retrospective study among 1000 patients with breast carcinomas and identified four cases of breast MEC. Clinical and demographic data were collected. Immunohistochemistry panels which were used to diagnose salivary gland MEC and breast carcinomas were also performed. MAML2 rearrangements were detected by FISH and fusion partners were identified by RNA sequencing. Whole-exome sequencing (WES) was used to reveal the genomes of these four breast MEC. Then, the biological functions and features of breast MEC were further compared with those of invasive breast carcinomas and salivary gland MEC.According to Ellis and Auclair’s methods, these four breast MEC could be classified as low-grade breast MEC. All the patients were alive, and disease-free survival (PFS) ranged from 20 months to 67 months. Among these four breast MEC, two cases were triple-negative, and the other two cases were found to be ER positive, with one also showing HER2 equivocal by immunohistochemical staining, but no amplification in FISH. FISH analysis confirmed the presence of the MAML2 translocation in three of four tumors, and CRTC1-MAML2 fusion was confirmed in two of them by RNA-sequencing. The average coverage size of WES for the tumor mutation burden estimation was 32 Mb. MUC4, RP1L1 and QRICH2 mutations were identified in at least three tumors, and these mutation also existed in breast invasive carcinoma databases (TCGA, Cell 2015; TCGA, Nature 2012). The results showed that there were many genes in breast MEC overlapping with the breast invasive carcinoma databases mentioned above, range from 5 to 63 genes (median:21 genes). Next, we assessed immune cell infiltration levels in these tumors. In all these tumors, M2 macrophages and plasma cell were in the high infiltration group. Our breast MEC showed different results from the salivary gland MEC, whose plasma cells were in the low infiltration group. Overall, we first analyzed the genomics and tumor microenvironment of breast mucoepidermoid carcinoma and proposed our hypothesis that although MECs arising in the breast resemble their salivary gland counterparts phenotypically, our findings indicate that breast MECs probably resemble invasive breast carcinomas at the genetic level and immune cell infiltration levels. More cases and in deep research need to be done to further understand this rare carcinoma.

Tile drainage as a driver of streamflow flashiness in agricultural areas of the Midwest, USA

AbstractThe Midwest of the USA is a highly productive agricultural region, in part due to the installation of perforated subsurface pipes, known as tile drains that remove excess water from wet soils. Tile drains rapidly move water to nearby streams, influencing how quickly streamflow rises and falls (i.e., streamflow “flashiness”). Currently, there are no comprehensive studies that compare the extent to which tile drainage influences flashiness across large and diverse agricultural regions. We address this knowledge gap by examining growing‐season (April–October) flashiness using the Richards‐Baker Index (RBI) in 139 watersheds located throughout the Midwest. Using a spatial tile‐drainage dataset, watersheds were split into low, medium, and high tile‐drainage classes. We found no significant differences between the flashiness of these three classes using a one‐way Kruskal–Wallis test. When watersheds were separated into infiltration groups to help control for different soil types, the high tile‐drainage class RBI was significantly higher than the low tile‐drainage class RBI in the high infiltration group. To further understand the causes of flashiness, additional environmental variables and their relationship to flashiness were examined using multivariate regression. In the low infiltration group, tile drainage significantly reduced flashiness, with watershed area and average depth to water table being the largest influences on flashiness. Tile drainage produced a larger reduction in flashiness in the high infiltration watersheds, with the largest influences being percent clay in the watershed and watershed area. These results indicate that the influence of tile drainage on flashiness emerges only after other watershed variables are accounted for. Given that tile drainage may increase in the future as precipitation patterns and extremes change, flashiness will likely continue to be modified. These results lead to an improved understanding of flood‐generating and nutrient transport mechanisms that are relevant to stakeholders across a wide range of sectors.

Immunohistochemical Expression of Programmed Death Ligand-1 and CD-8 in Glioblastoma

Background Glioblastoma (GBM) is the most common and most aggressive primary malignant brain tumor in adults. It is characterized by its extremely poor prognosis despite the available treatment strategies. Immune evasion is described in Glioblastoma via the PD-L1/ PD-1 interaction. Programmed death ligand 1 (PD-L1) is a cell surface ligand expressed on the surface of several tumors including GBM and its receptor is the PD-1 (Programmed deasth-1). Other malignant tumors including melanoma and non-small cell lung cancer, showed promising results using PD-L1/PD1 blockade treatment strategies where PD-L1 was described as a potential predictive biomarker for patient selection in these tumors. In our study we will shed the light on the expression and role of PD-L1 in glioblastoma, the presence of tumor infiltrating lymphocytes (TILs) and its interaction with PD-L1 in glioblastoma. The role of PD-L1, CD-8+ TILs, other clinicopathologic parameters and patient’s survival will be also discussed in relation to glioblastoma. Methods Paraffin embedded tissue blocks of thirty cases of GBN were retrieved, slides were prepared and stained via Hx&amp; E for evaluation and confirmation of diagnosis according to the CNS WHO 2016. Immunohistochemistry using PD-L1 and CD-8 was performed on the 30 glioblastoma cases. Interpretation of the results and correlation between the expression of both markers and the clinicopathologic parameters was also done. Furthermore, survival analysis was done to correlate overall survival and progression free survival of the glioblastoma cases to the expression of PD-L1 and CD-8 markers and also the clinicopathologic parameters available. Results Diffuse/fibrillary PD-L1 was expressed in all cases (30/30) with variable percentages (16.7% in ≤ 25% of cases 5/30, 33.3% in &amp;gt; 25%- ≤ 50% of cases10/30, 16.7% in &amp;gt; 50%- ≤ 75% of cases 5/30 and 33.3% in &amp;gt; 75% of cases 10/30). The mean value of Diffuse/fibrillary PD-L1 expression was 57.6%. Membranous PD-L1 was expressed in 6/30 of the cases. TILs were set into two groups according to Han et al 2014; low infiltration group 17/30 (56.7%) of cases and high infiltration group 13/30 (43.3%) of cases. Expression of CD-8 was 10% (median percentage) and 14.3% (mean percentage). PD-L1 and CD-8 correlation was statistically significant (P 0.001) where high PD-L1 expression is associated with low CD-8 expression. PD-L1 expression was associated with worse overall survival (0.001) and worse progression free survival (P 0.026). CD-8expression did not affect the outcome. Correlation of age, sex, tumor site and laterality to outcome were statistically insignificant. In the results of our study PD-L1 expression was the only independent factor that affected the prognosis according to Cox variate analysis. Conclusion The incidence of PD-L1 expression in GBM patients is robust. Higher expression of PD-L1 is associated with lower TILs expression and worse prognosis.

Integrated single cell and bulk sequencing analysis identifies tumor reactive CXCR6+ CD8 T cells as a predictor of immune infiltration and immunotherapy outcomes in hepatocellular carcinoma.

Various immune cell types in the tumor microenvironment (TME) of hepatocellular carcinoma (HCC) have been identified as important parameters associated with prognosis and responsiveness to immunotherapy. However, how various factors influence immune cell infiltration remains incompletely understood. Hence, we investigated the single cell multi-omics landscape of immune infiltration in HCC, particularly key gene and cell subsets that influence immune infiltration, thus potentially linking the immunotherapy response and immune cell infiltration. We grouped patients with HCC according to immune cell infiltration scores calculated by single sample gene set enrichment analysis (ssGSEA). Differential expression analysis, functional enrichment, clinical trait association, gene mutation analysis, tumor immune dysfunction and exclusion (TIDE) and prognostic model construction were used to investigate the immune infiltration landscape through multi-omics. Stepwise regression was further used to identify key genes regulating immune infiltration. Single cell analysis was performed to explore expression patterns of candidate genes and investigate associated cellular populations. Correlation analysis, ROC analysis, Immunotherapy cohorts were used to explore and confirm the role of key gene and cellular population in predicting immune infiltration state and immunotherapy response. Immunohistochemistry and multiplexed fluorescence staining were used to further validated our results. Patients with HCC were clustered into high and low immune infiltration groups. Mutations of CTNNB1 and TTN were significantly associated with immune infiltration and altered enrichment of cell populations in the TME. TIDE analysis demonstrated that T cell dysfunction and the T cell exclusion score were elevated in the high and low infiltration groups, respectively. Six risk genes and five risk immune cell types were identified and used to construct risk scores and a nomogram model. CXCR6 and LTA, identified by stepwise regression, were highly associated with immune infiltration. Single cell analysis revealed that LTA was expressed primarily in tumor infiltrating T lymphocytes and partial B lymphocytes, whereas CXCR6 was enriched predominantly in T and NK cells. Notably, CXCR6+ CD8 T cells were characterized as tumor enriched cells that may be potential predictors of high immune infiltration and the immune-checkpoint blockade response, and may serve as therapeutic targets. We constructed a comprehensive single cell and multi-omics landscape of immune infiltration in HCC, and delineated key genes and cellular populations regulating immune infiltration and immunotherapy response, thus providing insights into the mechanisms of immune infiltration and future therapeutic control.

Computational recognition of lncRNA signature of tumor-infiltrating activated CD4+ memory T cells with potential implications in prognosis and immunotherapy of lung adenocarcinoma.

e20519 Background: CD4 memory T cells are an important components of the tumour microenvironment and influence tumourigenesis and progression. However, it has not been determined whether lncRNAs in lung adenocarcinoma (LUAD) play a role in activated CD4+ memory T cells (CD4amT). Methods: The LUAD dataset acquired by The Cancer Genome Atlas was used as a training cohort (TCGA-LUAD). For the validation cohort, we used the combination of lung adenocarcinoma datasets from GENE EXPRESSION OMNIBUS database acquired 2 LUAD datasets, and set as the validation cohort (GEO-LUAD). Lung cancer immunotherapy cohorts have been published to test immune responses. The CIBERSORT algorithm was used to assess immune cell infiltration of tumours. Univariate Cox analysis was used to screen for prognostic CD4amT-related lncRNAs (CD4amTRL). LASSO analysis and multivariate Cox analysis were then performed to construct model for prognostic lncRNAs, the effects of which were assessed by Kaplan-Meier curves and subject operating characteristics (ROC). Patients were divided into high and low risk groups based on CD4amTRL signature. We have analyzed differences in enrichment pathways between the two groups and subsequently we investigated genes associated with the cancer-immune process, the immune microenvironment, immunotherapy responses and multiple anti-cancer drug sensitivity correlations. Results: In the two LUAD cohorts, compared with the high infiltration of CD4amT, those in the low infiltration group had better OS benefits (TCGA-LUAD: HR = 0.678; P = 0.028; GEO-LUAD: HR = 0.435; P &lt; 0.0001). We identified 6 prognostic CD4amTRL and constructed prediction models. cd4 CD4amTRL signature distinguished significantly different OS between the high-risk and low-risk groups of TCGA-LUAD (HR = 4.26). The AUCs predicting 1-, 3- and 5-year survival were 0.737, 0.715 and 0.722, respectively. In the GEO-LUAD dataset, the HR was 2.03, while the AUCs predicting 1-, 3- and 5-year survival were 0.719, 0.681 and 0.630, respectively. The risk score model independently predicted OS in LUAD patients and was associated with immune checkpoints. Numerous genes related to the cancer-immune process were highly correlated. In addition, the risk model was significantly associated with multiple immune microenvironment features (p &lt; 0.001). OS was significantly different between the two immunotherapy cohorts in the high- and low-risk groups for CD4amTRL signature (P &lt; 0.001). Conclusions: Our novel risk model developed based on CD4amTRL signatures can aid in predicting clinical prognosis and guiding treatment in patients with LUAD.

Tertiary lymphoid structure prevalence and prognostic value in cervical cancer.

e17521 Background: Recurrent or progressive cervical cancer have limited second-line treatment options. Response rates are often poor to second-line therapy (average response rate of 15%). Identification of factors which predict response to immunotherapy and targets to enhance the immune response are critically needed in cervical cancer. Chronic inflammation can initiate an immune response in non-secondary lymphoid organs (SLO) and form a Tertiary Lymphoid Structure (TLS). TLS is composed of immune cells clustered and organized and responsible for immune cell chemotaxis, which impacts cancer therapeutic response. Chemokine ligand 13 (CXCL13) is related to B cell attraction and TLS formation. Recent work from our group demonstrated human papilloma virus (HPV) positive head and neck squamous cell carcinoma (HNSCC) exhibited greater tumor infiltrating B cells (TIL-Bs) and TLS vs HPV negative disease indicating a role for viral infection in immune infiltration. Most cervical cancer is caused by HPV infection, therefore we investigated prognostic significance of immune infiltration in cervix cancer. Methods: A cohort analysis was conducted on 43 patients diagnosed with early stage cervical cancer. The presence of B cells, CD8 T cells, and CXCL13 was analyzed using singleplex immunohistochemistry staining. We separated infiltration into high infiltration and low infiltration, defined by their median value. TLS was identified using a multiplex immunofluorescence for TLS maturity panel. Histological findings were associated with cohort data. Results: High intratumoral infiltration of CD8 T cells was associated with longer overall survival in cancer patients. Median survival was 45 months for low infiltration group, whereas it was not reached by higher T cell infiltration (p &lt; 0.05). The prognostic value of T cell infiltration was stronger in adenocarcinoma, typically associated with worse outcomes, than in squamous cell carcinoma. In adenocarcinoma, median survival was 58 months for low T cell infiltration, it was not reached by high infiltration group. CXCL13 levels were prognostic for recurrence-free survival, with median survival of 53 months in low expression group and not reached in high CXCL13 presence group (p &lt; 0.05). The presence of TLS compared to low B cell infiltration was associated to higher survival, with 0% of deaths in the TLS group vs 40% in low B cell infiltration. While there was no correlation between TIL-B and patient outcomes, the presence of B cells in the aggregation process and higher CXCL13 levels were associated with improved survival, with 9% deaths vs 36% in low B cell group, possibly due to the support of TLS formation by B cell aggregation surrounded by CXCL13. Conclusions: Our study suggests that the presence of TLS, whether forming or established, is linked to improved clinical outcomes in cervical cancer. Further research is necessary to investigate the response of this cancer type to immunotherapy.

MP17-12 IMMUNE SCORE OF TUMOR-INFILTRATING CD8+ AND FOXP3+ LYMPHOCYTES AS A NOVEL PROGNOSTIC BIOMARKER IN HIGH-RISK LOCALIZED PROSTATE CANCER WHO UNDERWENT RADICAL PROSTATECTOMY

You have accessJournal of UrologyCME1 Apr 2023MP17-12 IMMUNE SCORE OF TUMOR-INFILTRATING CD8+ AND FOXP3+ LYMPHOCYTES AS A NOVEL PROGNOSTIC BIOMARKER IN HIGH-RISK LOCALIZED PROSTATE CANCER WHO UNDERWENT RADICAL PROSTATECTOMY Hiromi Sato, Shintaro Narita, Yoshiko Takahashi, Masanori Ishida, Mizuki Kobayashi, Soki Kasima, Ryohei Yamamoto, Taketoshi Nara, Mingguo Huang, Kazuyuki Numakura, Mitsuru Saito, Hiroshi Nanjo, and Tomonori Habuchi Hiromi SatoHiromi Sato More articles by this author , Shintaro NaritaShintaro Narita More articles by this author , Yoshiko TakahashiYoshiko Takahashi More articles by this author , Masanori IshidaMasanori Ishida More articles by this author , Mizuki KobayashiMizuki Kobayashi More articles by this author , Soki KasimaSoki Kasima More articles by this author , Ryohei YamamotoRyohei Yamamoto More articles by this author , Taketoshi NaraTaketoshi Nara More articles by this author , Mingguo HuangMingguo Huang More articles by this author , Kazuyuki NumakuraKazuyuki Numakura More articles by this author , Mitsuru SaitoMitsuru Saito More articles by this author , Hiroshi NanjoHiroshi Nanjo More articles by this author , and Tomonori HabuchiTomonori Habuchi More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003237.12AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: For several cancer patients, immunotherapy has resulted in better outcomes, and tumor-infiltrating lymphocytes (TILs) have promising potential to be associated with prognosis of cancer. However, the clinical significance of TILs in high-risk localized prostate cancer (HRLPCa) is largely unknown. Here we investigate the effect of tissue-resident T cells on clinical outcomes in patients with HRLPCa who had a radical prostatectomy (RP). METHODS: Tumor microarrays were built using prostatectomy specimens from 240 cores of 80 HRLPCa treated with RP. Each specimen was scored individually using immunohistochemistry for the mean numbers of the CD3+, CD4+, CD8+, and FoxP3+ TILs (IHC). After being classified into high- and low-infiltration groups for each TILs, the association between the immune scores, clinicopathological characteristics, and biochemical recurrence-free survival (bRFS) was statistically evaluated. RESULTS: Patients in the higher infiltrate groups for CD3+, CD4+, CD8+, and FoxP3+ TILs had significantly lower PSA levels than those in the lower infiltrate groups, and the rate of pathological T3–4 disease was significantly higher in the low CD4+ group than in the CD4+ high group (p=0.015). No significant difference in bRFS was observed between the low and high groups of each TILs. The FoxP3 high+CD8 low group had significantly worse bRFS than the other groups (2-year bRFS: 88.5% vs. 60.0%, p=0.004) after the groups were divided into four groups, including FoxP3 low +CD8 low, FoxP3 low+CD8 high, FoxP3 high+CD8 low, and FoxP3 high+CD8 high. In contrast, the combination of CD4 and FoxP3 scores was not statistically related to bPFS (p=0.228). FoxP3 high+CD8 low was found to be an independent predictor of bRFS outcome in the multivariate analysis (hazard ratio: 3.431; 95% CI: 1.048–11.240; p=0.049); as well as pathological T3–4 diseases (hazard ratio: 3.546; 95% CI: 1.250–10.060; p=0.017). CONCLUSIONS: The combined IHC scores with CD8 and FoxP3 in RP specimens may be a potential biomarker for biochemical recurrence in patients with HRLPCa. Source of Funding: GlaxoSmithKline plc © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e217 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Hiromi Sato More articles by this author Shintaro Narita More articles by this author Yoshiko Takahashi More articles by this author Masanori Ishida More articles by this author Mizuki Kobayashi More articles by this author Soki Kasima More articles by this author Ryohei Yamamoto More articles by this author Taketoshi Nara More articles by this author Mingguo Huang More articles by this author Kazuyuki Numakura More articles by this author Mitsuru Saito More articles by this author Hiroshi Nanjo More articles by this author Tomonori Habuchi More articles by this author Expand All Advertisement PDF downloadLoading ...

A novel immune cell signature for predicting osteosarcoma prognosis and guiding therapy.

Dysregulation of immune cell infiltration in the tumor microenvironment contributes to the progression of osteosarcoma (OS). In the present study, we explored genes related to immune cell infiltration and constructed a risk model to predict the prognosis of and guide therapeutic strategies for OS. The gene expression profile of OS was obtained from TARGET and Gene Expression Omnibus, which were set as the discovery and verification cohorts. CIBERSORT and Kaplan survival analyses were used to analyze the effects of immune cells on the overall survival rates of OS in the discovery cohort. Differentially expressed gene (DEG) analysis and protein–protein interaction (PPI) networks were used to analyze genes associated with immune cell infiltration. Cox regression analysis was used to select key genes to construct a risk model that classified OS tissues into high- and low-risk groups. The prognostic value of the risk model for survival and metastasis was analyzed by Kaplan–Meier survival analyses, receiver operating characteristic curves, and immunohistochemical experiments. Immunological characteristics and response effects of immune checkpoint blockade (ICB) therapy in OS tissues were analyzed using the ESTIMATE and Tumor Immune Dysfunction and Exclusion algorithms, while sensitivity for both targeted and chemotherapy drugs was analyzed using the OncoPredict algorithm. It was demonstrated that the high infiltration of resting dendritic cells in OS tissues was associated with poor prognosis. A total of 225 DEGs were found between the high- and low-infiltration groups of OS tissues, while 94 genes interacted with others. Through COX analyses, among these 94 genes, four genes (including AOC3, CDK6, COL22A1, and RNASE6) were used to construct a risk model. This risk model showed a remarkable prognostic value for survival rates and metastasis in both the discovery and verification cohorts. Even though a high microsatellite instability score was observed in the high-risk group, the ICB response in the high-risk group was poor. Furthermore, using OncoPredict, we found that the high-risk group OS tissues were resistant to seven drugs and sensitive to 25 drugs. Therefore, our study indicates that the resting dendritic cell signature constructed by AOC3, CDK6, COL22A1, and RNASE6 may contribute to predicting osteosarcoma prognosis and thus therapy guidance.

Identification and Validation of Immune Infiltration Phenotypes in Laryngeal Squamous Cell Carcinoma by Integrative Multi-Omics Analysis

BackgroundLaryngeal squamous cell carcinoma (LSCC) is one of the world’s most common head and neck cancer. However, the immune infiltration phenotypes of LSCC have not been well investigated.MethodsThe multi-omics data of LSCC were obtained from the TCGA (n=111) and GEO (n=57) datasets. The infiltrations of the 24 immune cell populations were calculated using the GSVA method. Then LSCC samples with different immune cell infiltrating patterns were clustered, and the multi-omics differences were investigated.ResultsPatients were clustered into the high-infiltration and low-infiltration groups. The infiltration scores of most immune cells were higher in the high-infiltration group. Patients with high-infiltration phenotype have high N and TNM stages but better survival, as well as less mutated COL11A1 and MUC17. Common targets of immunotherapies such as PD1, PDL1, LAG3, and CTLA4 were significantly up-regulated in the high-infiltration group. The differentially expressed genes were mainly enriched in several immune-related GOs and KEGG pathways. Based on the genes, miRNAs, and lncRNAs differentially expressed in both the TCGA and GEO cohorts, we built a ceRNA network, in which BTN3A1, CCR1, miR-149-5p, and so on, located at the center. A predictive model was also constructed to calculate a patient’s immune infiltration phenotype using 16 genes’ expression values, showing excellent accuracy and specificity in the TCGA and GEO cohorts.ConclusionsIn this study, the immune infiltration phenotypes of LSCC and the corresponding multi-omics differences were explored. Our model might be valuable to predicting immunotherapy’s outcome.

The Combined Model of CX3CR1-Related Immune Infiltration Genes to Evaluate the Prognosis of Idiopathic Pulmonary Fibrosis.

BackgroundHigh expression of chemokine (C-X3-C motif) receptor 1 (CX3CR1) was shown to contribute to the progression of many fibrotic diseases. However, there is still no study for the role of CX3CR1 in idiopathic pulmonary fibrosis (IPF). Therefore, we aimed to identify CX3CR1-related immune infiltration genes (IIGs) in IPF and establish a combined risk model to evaluate the prognosis of IPF.MethodsA discovery cohort of IPF patients (GSE70867) was downloaded from the Gene Expression Omnibus dataset. We identified the composition of 22 kinds of immune cells infiltration by CIBERSORT. The Cox regression model with the LASSO method was used for identifying prognostic genes and developing CX3CR1-related IIGs. Kaplan–Meier was applied to plot the survival curve of prognosis model. Peripheral blood mononuclear cell (PBMC) and bronchoalveolar lavage fluid (BALF) were collected to be tested by quantitative reverse transcriptase-PCR (qRT-PCR) from 15 clinical samples, including 8 healthy controls (HC), 4 patients with usual interstitial pneumonia (UIP) and 3 patients with pulmonary fibrosis (FIB).ResultsWe found that high expression of CX3CR1 in BALF contributed to the poor prognosis in IPF patients. ALR4C, RAB37, GPR56, MARCKS, PXN and RASSF2 were identified as CX3CR1-related IIGs, which were highly expressed in PBMC of UIP/FIB patients than that of HC. Moreover, the expression of PXN was higher in FIB patients’ PBMC than that of UIP ones. In the cohort of IPF patients, high infiltration of activated NK cells in BALF caused poor survival compared to low infiltration group. The infiltration of activated NK was regulated by CX3CR1-related IIGs. The combined risk model predicted that high expression of CX3CR1-related IIGs and high infiltrated activated NK cells caused poor prognosis in IPF patients.ConclusionWe identified a group of CX3CR1-related IIGs in IPF patients. This combined risk model provided new insights in the prognosis and therapy of IPF.

Characteristics of CD8+ T cell infiltration in colorectal cancer and their correlation with prognosis

Objective: As cytotoxic T cells, CD8+ T lymphocytes can kill tumor cells by releasing perforin and other effector molecules, but the correlation between their infiltration level and the prognosis of colorectal cancer varies in previous studies. This study aims to explore the distribution of CD8+T cells in tumor center and invasive margin of colorectal cancer, and to analyze their correlation with the prognosis of patients. Methods: A retrospective cohort study was used to analyze the clinicopathological features of 221 patients with colorectal cancer from the colorectal cancer pathological database of the Sixth Affiliated Hospital of Sun Yat-sen University between 2009 and 2012. Case inclusion criteria: (1) colorectal cancers confirmed by postoperative pathology; (2) patients with follow-up data. Exclusion criteria: (1) multiple primary cancers; (2) inflammatory bowel disease, Lynch syndrome or familial adenomatous polyposis; (3) no available paraffin slides; (4) patients receiving preoperative radiotherapy or chemotherapy. A total of 221 patients met the criteria. Immunohistochemical staining was used to count the CD8+ T cells in tumor center and invasive margin in the paraffin slides. Meanwhile the relative expression of CD8B gene in 22 fresh freeze samples of colorectal cancer was detected. Then the correlation of the expression with CD8+T cell density was examined. The patients were divided into high and low infiltration groups according to the level of CD8+T cells. Log-rank test was applied to compare the overall survival of the two groups of patients, and Cox regression analysis was used to adjust the prognostic significance of CD8+T cell infiltration. Results: There were 118 males and 103 females. In 221 slides, CD8+T cells infiltrating in invasive margin were more than those in tumor center [median (range): 37(0-141) / field vs. 14(0-106) / field, Z=-11.985, P<0.001], and the number of CD8+T cell in the tumor center was positively correlated with those in invasive margin (r=0.610, P<0.001). The number of CD8+ T cell in tumor center was positively correlated with the relative expression of CD8B gene (r=0.524, P=0.012). Survival analysis showed that the overall survival of the high infiltration group was better than that of the low infiltration group both in tumor center and invasive margin (median survival: 84.1 months vs. 73.5 months, P<0.001; 84.2 months vs. 75.9 months, P=0.002). Cox regression analysis revealed that high CD8+T cell infiltration in tumor center was an independent protective factor of overall survival (HR=0.369, 95% CI: 0.168-0.812, P=0.013). Conclusions: The infiltration level of CD8+T cells in tumor center is lower than that in invasive margin, and they are positively correlated. The level of CD8+ T cell infiltration in tumor center is related to overall survival and can be used as a potential pronostic marker.

Heterogeneity analysis of the immune microenvironment in laryngeal carcinoma revealed potential prognostic biomarkers.

Laryngeal squamous cell cancer (LSCC) is the second most prevalent malignancy occurring in the head and neck with a high incidence and mortality rate. Immunotherapy has recently become an emerging treatment for cancer. It is therefore essential to explore the role of tumour immunity in laryngeal cancer. Our study first delineated and evaluated the comprehensive immune infiltration landscapes of the tumour microenvironment in LSCC. A hierarchical clustering method was applied to classify the LSCC samples into two groups (high- and low-infiltration groups). We found that individuals with low immune infiltration characteristics had significantly better survival than those in the high-infiltration group, possibly because of the elevated infiltration of immune suppressive cells, such as regulatory T cells and myeloid-derived suppressor cells, in the high-infiltration group. Differentially expressed genes between two groups were involved in some immune-related terms, such as antigen processing and presentation. A univariate Cox analysis and least absolute shrinkage and selection operator analysis were performed to identify an immune gene-set-based prognostic signature (IBPS) to assess the risk of LSCC. The prognostic model comprising six IBPSs was successfully verified to be robust in different cohorts. The expression of the six IBPSs was detected by immunohistochemistry in 110 cases of LSCC. In addition, different inflammatory profiles and immune checkpoint landscape of LSCC were found between two groups. Hence, our model could serve as a candidate immunotherapeutic biomarker and potential therapeutic target for laryngeal cancer.

Macrophages M1-Related Prognostic Signature in Hepatocellular Carcinoma.

A large number of studies have found that macrophages M1 play an important role in the occurrence and development of tumors. The aim of our study is to explore the causes of differential infiltration of macrophages M1 in hepatocellular carcinoma from the perspective of transcriptome and establish a prognostic model of hepatocellular carcinoma. We downloaded gene expression and clinical data from the public database, estimated the content of macrophages M1 in different samples with R software, and found the different genes between high- and low-infiltration groups. Using differentially expressed genes, we constructed a model composed of 7 genes. The risk score of the model has a good ability to predict the prognosis, has a positive correlation with immune checkpoints, and is closely related to other immune cells and immune function. Our model shows good prognostic function and has wide application value.

Identification and validation of a novel immune-related signature associated with macrophages and CD8 T cell infiltration predicting overall survival for hepatocellular carcinoma

BackgroundAlthough the effects of macrophages and CD8 T cell infiltration on clinical outcome have been widely reported, the association between immunity-associated gene with them for hepatocellular carcinoma (HCC) remains unclear.Materials and methodsThe ssGSEA served for quantifying the macrophages as well as CD8 T cell infiltration in the HCC samples obtained from TCGA database. Kaplan–Meier (KM) survival assay was used to determine the associations between macrophages and CD8 T cell infiltration with OS. LASSO Cox regressive method assisted in developing an immune gene signature as well as building a risk score. The performance was evaluated by the time-dependent ROC together with the KM survival analysis. The ICGC database were adopted for external verification. CIBERSORT was applied to the correlation analysis on the immune-related signature and the immunocyte infiltration. GSEA were employed exploring the underlying molecular mechanisms.ResultsIncreased CD8+ T cell infiltration was associated with longer OS, whereas a greater infiltration of macrophages was related to shorter OS. There were 398 differential expression genes (DEGs) between the high- and low infiltration groups with the “edgeR” package. An prognostic signature consisted of 10 immune genes was built in TCGA and examined in ICGC. The uniform cutoff (0.927) was adopted for separating sufferers into the high-risk (HR) and low-risk (LR) groups. The ROC curves revealed that the AUC data for this signature predicting 1, 2, 3, 4 and 5 year were all above 0.7 in both TCGA and ICGC cohort and patients in the HR group exhibited an evidently weaker prognostic results compared with the LR group. The HR group presented evidently greater Tregs and Macrophage M0 relative to the LR group, whereas the LR group saw the enrichment of CD8 T cells.ConclusionThe immune signature associated with macrophages as well as CD8 T cell infiltration has reliable prognostic and predictive value for HCC patients.

Identification and validation of tumor microenvironment-related lncRNA prognostic signature for uveal melanoma.

To investigate the role of tumor microenvironment (TME)-related long non-coding RNA (lncRNA) in uveal melanoma (UM), probable prognostic signature and potential small molecule drugs using bioinformatics analysis. UM expression profile data were downloaded from the Cancer Genome Atlas (TCGA) and bioinformatics methods were used to find prognostic lncRNAs related to UM immune cell infiltration. The gene expression profile data of 80 TCGA specimens were analyzed using the single sample Gene Set Enrichment Analysis (ssGSEA) method, and the immune cell infiltration of a single specimen was evaluated. Finally, the specimens were divided into high and low infiltration groups. The differential expression between the two groups was analyzed using the R package 'edgeR'. Univariate, multivariate and Least Absolute Shrinkage and Selection Operator (LASSO) Cox regression analyses were performed to explore the prognostic value of TME-related lncRNAs. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional analyses were also performed. The Connectivity Map (CMap) data set was used to screen molecular drugs that may treat UM. A total of 2393 differentially expressed genes were identified and met the criteria for the low and high immune cell infiltration groups. Univariate Cox analysis of lncRNA genes with differential expression identified 186 genes associated with prognosis. Eight prognostic markers of TME-included lncRNA genes were established as potentially independent prognostic elements. Among 269 differentially expressed lncRNAs, 69 were up-regulated and 200 were down-regulated. Univariate Cox regression analysis of the risk indicators and clinical characteristics of the 8 lncRNA gene constructs showed that age, TNM stage, tumor base diameter, and low and high risk indices had significant prognostic value. We screened the potential small-molecule drugs for UM, including W-13, AH-6809 and Imatinib. The prognostic markers identified in this study are reliable biomarkers of UM. This study expands our current understanding of the role of TME-related lncRNAs in UM genesis, which may lay the foundations for future treatment of this disease.

Immune Landscape of Invasive Ductal Carcinoma Tumor Microenvironment Identifies a Prognostic and Immunotherapeutically Relevant Gene Signature.

Background: Invasive ductal carcinoma (IDC) is a clinically and molecularly distinct disease. Tumor microenvironment (TME) immune phenotypes play crucial roles in predicting clinical outcomes and therapeutic efficacy.Method: In this study, we depict the immune landscape of IDC by using transcriptome profiling and clinical characteristics retrieved from The Cancer Genome Atlas (TCGA) data portal. Immune cell infiltration was evaluated via single-sample gene set enrichment (ssGSEA) analysis and systematically correlated with genomic characteristics and clinicopathological features of IDC patients. Furthermore, an immune signature was constructed using the least absolute shrinkage and selection operator (LASSO) Cox regression algorithm. A random forest algorithm was applied to identify the most important somatic gene mutations associated with the constructed immune signature. A nomogram that integrated clinicopathological features with the immune signature to predict survival probability was constructed by multivariate Cox regression.Results: The IDC were clustered into low immune infiltration, intermediate immune infiltration, and high immune infiltration by the immune landscape. The high infiltration group had a favorable survival probability compared with that of the low infiltration group. The low-risk score subtype identified by the immune signature was characterized by T cell-mediated immune activation. Additionally, activation of the interferon-α response, interferon-γ response, and TNF-α signaling via the NFκB pathway was observed in the low-risk score subtype, which indicated T cell activation and may be responsible for significantly favorable outcomes in IDC patients. A random forest algorithm identified the most important somatic gene mutations associated with the constructed immune signature. Furthermore, a nomogram that integrated clinicopathological features with the immune signature to predict survival probability was constructed, revealing that the immune signature was an independent prognostic biomarker. Finally, the relationship of VEGFA, PD1, PDL-1, and CTLA-4 expression with the immune infiltration landscape and the immune signature was analyzed to interpret the responses of IDC patients to immunotherapy.Conclusion: Taken together, we performed a comprehensive evaluation of the immune landscape of IDC and constructed an immune signature related to the immune landscape. This analysis of TME immune infiltration landscape has shed light on how IDC respond to immunotherapy and may guide the development of novel drug combination strategies.

The Clinical Significance of Tumor Associated Macrophages in Myelodysplastic Syndromes

Objective:The purpose of this study is to investigate the relationship between the infiltration of tumor associated macrophages (TAMs) and the disease risk stratification and survival of patients with myelodysplastic syndrome (MDS) and to explore the role of TAMs in the clinical prognosis.Methods:We retrospectively collected and analyzed 115 patients initially diagnosed wih myelodysplastic syndromes from January, 2010 to July, 2017 in West China Hospital of Sichuan University. Both bone marrow biopsy specimens and clinical data of the patients enrolled were collected. All patients were assessed prognosis by international prognostic scoring system (IPSS). We quantified the involvement of macrophage (MΦ), alternatively activated macrophage (M2 MΦ) and classic activated macrophage (M1 MΦ) in bone marrow specimen by staining with anti-CD68 monoclonal antibody, anti-CD163 monoclonal antibody, and anti-iNOS monoclonal antibody respectively. Log-rank test was used to evaluate the difference of overall survival (OS) among different subgroups. Logistic regression was used to evaluate the effect of clinical parameters on patients' OS. Cox proportional-hazards models were used to estimate the independent risk factors influencing the prognosis of patients.Results:1 In the high risk group of MDS, the composition of CD163+MΦ was higher than that of the low risk group(p < 0.001). In the low risk group of MDS, the composition of iNOS+ MΦ was higher than that of the group with high risk(p < 0.001).2 The median survival time and 3-year survival rate in the high CD163+MΦinfiltration group were significantly lower than those in the low infiltration group (13 months vs 43 months, 27% vs 64%, Log-rank test, P<0.001). The median survival time and 3-year survival rate in high iNOS+MΦ infiltration group were significantly higher than those in low infiltration group (42. 5 months vs 11 months, 65% vs 27% Log-rank test, P< 0.001).3 According to the infiltration degree of iNOS+MΦ and CD163+MΦ, the M1 (iNOShigh/CD 163low)and M2 TA(iNOSlow/CD163high) MΦ were further distinguished (The median OS were 53m and 15m respectively, p< 0.001).4 In multivariate analysis, high infiltration of CD163+MΦ and CD68+MΦ, low infiltration of iNOS+MΦ and low PLT level were independent risk factors for patients' OS .Conclusion and Discuss:The degree of bone marrow TAMs subtype infiltration in MDS patients was associated with disease risk group. CD163+M Φ was the main TAMs in the high risk group of MDS,while iNOS+M Φ was the main TAMs in the low risk group. There was a positive correlation between the infiltration of M1-TAMs and OS. On the contrary, there was a negative correlation between the infiltration of M2-TAMs and OS. Analyzing TAMs subsets in bone marrow of patients with MDS is helpful to evaluate disease risk and prognosis. DisclosuresNo relevant conflicts of interest to declare.

Neutrophils in primary gastric tumors are correlated with neutrophil infiltration in tumor-draining lymph nodes and the systemic inflammatory response

BackgroundTumor-Associated Neutrophils (TANs) may be able to induce lymphangiogenesis and angiogenesis, although the detailed roles of TANs remain unclear. The Neutrophil-Lymphocyte Ratio (NLR) is an inflammation-based prognostic factor for gastric cancer. This study aimed to investigate the distribution of CD15+neutrophils in the primary tumor and Tumor-Draining Lymph Nodes (TDLNs), and to examine the association of TANs with the clinicopathological features (including NLR) of patients with gastric cancer.ResultsImmunohistochemical staining showed that the median number of CD15+TANs was 18 and 24 per high-power field (HPF) in primary tumors and TDLNs, respectively. Patients were divided into high and low infiltration groups based on the median number. A high number of infiltrating CD15+TANs in the primary tumors and in the TDLNs were associated with depth of invasion and lymph node metastasis. Kaplan-Meier analysis revealed that a poor overall survival was associated with high numbers of CD15+TANs, and the multivariate analyses revealed that a high number of CD15+TANs in the TDLNs was an independent prognostic factor. The numbers of CD15+TANs in the primary tumors and TDLNs showed weak positive correlation. The number of CD15+TANs in the primary tumors was positively correlated with the preoperative NLR, (P = 0.001, R = 0.327) and immunohistochemical staining revealed that C-X-C motif chemokine receptor 2 (CXCR2) +neutrophils might be the origin of the CD15+TANs. Flow cytometry analysis indicated that infiltrating neutrophils increased in the tumor and TDLN compared to non-cancerous tissue. Neutrophils treated with cancer supernatant upregulated TWIST and IL-6 genes in vitro.ConclusionOur findings suggested that local infiltration of CD15+TANs may be correlated with inflammation in TDLNs and systemic response to cause metastasis in gastric carcinoma.

Association of tumor infiltrating CD8 positive T cells and PDL-1 expression with survival after conversion surgery for unresectable gastric cancer.

62 Background: Conversion surgery after chemotherapy responds to unresectable gastric cancer has long-term prognosis, however, chemotherapy after surgery should also be considered to improve treatment outcome. The effectiveness of immune checkpoint inhibitors for gastric cancer has been proved. In this study, we investigated the association between PDL-1 expression and CD8+ T cell infiltration in the primary tumor and prognosis of patients who underwent conversion surgery after S-1-based multidrug chemotherapy in unresectable gastric cancer. Methods: We evaluated PDL-1 expression and CD8 T cell infiltration by immunohistochemical staining and examined the relationship with prognosis in 90 patients with Stage IV gastric cancer who underwent gastrectomy in our department from 2007 to 2015. Results: 90 cases included 47 cases of palliative resection, 26 cases of CY1P 0cases and 17 cases of conversion surgery. Among Conversion Surgery cases, 11 (65%) patients underwent R0 surgery, and the 5-year survival rate was 66%. The 5-year survival rate of non-curative resected cases was 16%. In immunohistochemical examination, PDL-1 high expression was a poor prognostic factor in palliative surgery and CY1P0 cases. Meanwhile, in the case of Conversion Surgery, the 5-year survival rate was 58%, 32%, 61%, and 30% in the order of PDL-1 high expression group, low expression group, CD8 T cell high infiltration, and low infiltration group, respectively. Prognosis of PDL-1 high expression group and CD8 T cell infiltration group was good. There was no association between curative resection and neither of CD8 T cell infiltration or PDL-1 expression. Conclusions: We found that the prognosis was relatively good in patients where local immune response was enhanced, regardless of R0 excision. Our results suggested that infiltration of CD8 T cells in the primary tumor may be a prognostic predictor, and it is expected to use an immune check point inhibitor after conversion surgery for unresectable gastric cancer.