Regulation Of Tumor Immunity Research Articles

Abstract 4188: CD4+ T cells mediate non-canonical rejection of major histocompatibility class-I deficient pancreatic tumors independently of CD8+ T cells

Abstract Identifying and interrogating novel immune-mediated mechanisms of tumor rejection is a major clinical opportunity. Immune checkpoint blockade (ICB) induces tumor rejection primarily via cytolytic CD8+ T cells, but many patients have primary or acquired resistance to CD8+ T cell immunity. Reduced major histocompatibility complex I (MHC I) expression is a contributing factor to resistance, and pancreatic ductal adenocarcinoma (PDAC) is historically treatment-refractory due to, among other features, low MHC expression and a dearth of T cells in the tumor. Interrogating interventions in the context of the PDAC tumor microenvironment will reveal novel mechanisms regulating sensitivity to immune-mediated destruction. Using a genetically engineered mouse model of PDAC, we previously reported that combination immunotherapy (agonistic anti-CD40 and dual ICB, anti-PD-1/CTLA-4) induced both a CD8+ and a CD4+ T cell response sufficient to mediate tumor rejection and long-lived protection. Here, we utilized CRISPR-Cas9 to disrupt the dominant MHC I (H-2Kb allele) expression in PDAC cell lines, precluding direct antigen presentation to CD8+ T cells via this molecule. Subcutaneous injection of MHC I-deficient tumor clones into wild-type mice revealed normal tumor growth at baseline, and significantly delayed tumor growth kinetics in response to combination immunotherapy (CD40/ICB) in a CD4+ T cell-dependent manner (p < 0.0001 vs. vehicle-treated mice). These findings were observed in parental PDAC clones lacking H-2Kb expression, as well as in an ovalbumin (OVA)-expressing PDAC clone. CD4+ T cells mediated tumor rejection including in OVA-tolerized mice, excluding the role of tumor-neoantigens as a major determinant of response. There was no direct contribution of tumor rejection by CD8+ T cells, despite the generation of an activated, tumor-specific CD8+ T cell response in the tumor site (24.5% vs. 1.98% in vehicle-treated mice, p < 0.003). CD4+ T cells upregulated IFN-γ production (52.3% vs. 11.9% in vehicle-treated mice, p < 0.0001), and host IFN-γ and IFN-γ receptor expression were required for response. No changes were observed in the expression of cytotoxic molecules (Granzyme A or B) by CD4+ T cells after CD40/ICB, and perforin was not required for tumor rejection. Furthermore, depletion of macrophage and myeloid cells or genetic knockout of inducible nitric oxide synthase did not dampen CD4+ T cell immunity against established PDAC tumors. Thus, CD4+ T cells mediated PDAC tumor rejection via non-canonical mechanisms, excluding cytotoxic molecules and re-direction of anti-tumor macrophage effector functions after treatment with anti-CD40/ICB. This study provides critical insight into novel mechanisms of tumor clearance beyond direct CD8+ T cell:tumor cell interactions, and opportunities to exploit CD4 T+ cells as regulators of tumor immunity. Citation Format: Samuel I. Kim, Charu Arora, Ioannis I. Verginadis, Christopher R. Cassella, Nune Markosyan, Constantinos Koumenis, Robert H. Vonderheide, Katelyn T. Byrne. CD4+ T cells mediate non-canonical rejection of major histocompatibility class-I deficient pancreatic tumors independently of CD8+ T cells [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 4188.

Abstract 4150: Enriching for adenosine antagonist patient responses through deep learning

Abstract Background: Adenosine signaling is a key metabolic pathway regulating tumor immunity. The conversion of inflammatory extracellular ATP into immunosuppressive adenosine invokes signaling of the A2a receptor (A2AR) in the tumor microenvironment. This dampens immune responses and creates a pro-tumor niche. A number of novel IO drugs targeting the adenosine pathway through inhibition of the ectonucleotidases CD39 and CD73 or the A2AR/BR are in clinical trials, including our non-CNS penetrant A2AR-selective antagonist, EXS21546 (NCT04727138). While early clinical results from other adenosine receptor-inhibitors (A2ARi) have shown modest monotherapy activity in nonspecific patient populations, we believe greater success can be achieved by leveraging methods that enable the evaluation of single-cell effects in patient samples preclinically. Using our deep learning driven image analysis platform, we define an adenosine-induced, tumor protective immunosuppression biomarker to augment A2AR antagonist responder identification. Here we describe efforts to transcriptionally and functionally map the adenosine suppressed immune potential and activation by inhibition of A2AR with EXS21546 in primary material. The goal is to reveal gene signatures indicative of adenosine immunosuppression deployable in clinical studies, increasing the likelihood of trial success by identifying patients that have the highest efficiency potential for A2AR targeted therapy. Methods: Leveraging patient material as disease relevant model systems, and collecting baseline and treatment condition transcriptomics, we model the patient specific anti-cancer immune repertoire and validate patient selection methods functionally with a translatable high content imaging platform amenable to primary human material supported by end-to-end deep learning driven image analysis. Results: Combining single cell transcriptomic and functional response data, we demonstrate preclinical mechanistic studies of A2AR antagonism on infiltrating immune cells, with the ultimate aim of discovering predictive algorithms to enrich patients more likely respond to adenosine pathway inhibitors. Patient selection gene signatures are functionally validated using a high content imaging platform with proven translational capabilities (Kornauth et al, Cancer Disc., 2021), demonstrating the association of anti-cancer immune activity with inhibition of adenosine signaling by EXS21546. Signatures and patient selection algorithms are cross-validated with publicly available data. Discussion: Gathering multiple layers of data from primary tumor tissues, we reveal and map the association of immune response potential to A2AR inhibition in cancer. Patient stratification gene signatures identified have the aim to be implemented in future studies of our candidate A2ARi EXS21546, to deliver the right drug at the right time in the right patients. Citation Format: Isabella Alt, Robert Shelke, Anna Lobley, Claudia Baumgaertler, Maja Stulic, Pierre Fons, Mark Whittaker, Klaus Hackner, Lucia Dzurillova, Edga Petru, Laudia Hadjari, Judith Lafleur, Josef Singer, Nikolaus Krall, Lukas Hefler, Thorsten Füreder, Christina Taubert, Christophe Boudesco, Andrew Payne, Gregory Ian Vladimer. Enriching for adenosine antagonist patient responses through deep learning [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 4150.

Development of a Platelet-Related Prognostic Model for Colorectal Cancer.

Colorectal cancer (CRC) represents one of the most common malignancies with high morbidity worldwide. Growing evidence has suggested that platelets are a fundamental component of the tumor microenvironment and play crucial roles in driving tumor biological behavior. The construction of a platelet-related prognostic model that can reliably predict CRC prognosis is of great clinical significance. The 1427 CRC-specific platelet-related genes were collected and mainly enriched in the ribosome and immune-related pathways. Based on platelet-related genes, three subtypes of TCGA CRC samples were identified by consensus clustering and characterized by differences in angiogenesis, epithelial–mesenchymal transition, immune infiltration, and prognosis. A total of 100 prognostic platelet-related genes were identified by univariate Cox regression. LASSO Cox regression further shrank those genes and constructed a 10-gene prognostic model. The patients with higher risk scores had significantly worse disease-specific survival than those with lower scores in both TCGA and validation cohorts. The risk score demonstrated good predictive performance for prognosis by receiver operating characteristic (ROC) curves. Furthermore, multivariate Cox regression analysis showed that the risk score was independent of TNM stage, sex, and age, and a graphic nomogram based on the risk score and clinical factors was developed to predict survival probability of CRC patients. Patients from the high-risk group were characterized by higher infiltration of immunosuppressive cells such as MDSC and Treg and higher expression of checkpoints CTLA4, CD86, and PDCD1LG2. Taken together, we identified three platelet-related subtypes and specifically constructed a promising 10-gene prognostic model in CRC. Our results highlighted the potential survival effects of platelet-related genes and provided evidence about their roles in regulating tumor immunity.

Mesothelial cell-derived antigen-presenting cancer-associated fibroblasts induce expansion of regulatory Tcells in pancreatic cancer.

Recent studies have identified a unique cancer-associated fibroblast (CAF) population termed antigen-presenting CAFs (apCAFs), characterized by the expression of major histocompatibility complex class II molecules, suggesting a function in regulating tumor immunity. Here, by integrating multiple single-cell RNA-sequencing studies and performing robust lineage-tracing assays, we find that apCAFs are derived from mesothelial cells. During pancreatic cancer progression, mesothelial cells form apCAFs by downregulating mesothelial features and gaining fibroblastic features, a process induced by interleukin-1 and transforming growth factor β. apCAFs directly ligate and induce naive CD4+ Tcells into regulatory Tcells (Tregs) in an antigen-specific manner. Moreover, treatment with an antibody targeting the mesothelial cell marker mesothelin can effectively inhibit mesothelial cell to apCAF transition and Treg formation induced by apCAFs. Taken together, our study elucidates how mesothelial cells may contribute to immune evasion in pancreatic cancer and provides insight on strategies to enhance cancer immune therapy.

Cytokine as an potential prognostic factor associated with absolute value of neutrophil and serum albumin in breast cancer patients.

e13007 Background: Cytokines are involved in many inflammatory diseases through various ways and play an important role in tumor immune regulation. With the development of molecular biology these years, researchers have found that breast cancer is not only related to genetic and environmental factors, but also to the chronic inflammation and immunity. But at the moment，there are few studies on the prognostic evaluation of serum cytokines in breast cancer. The correlation between serum cytokines and leukocytes was not clear. The purpose of this study is to investigate the value of serum cytokines in prognosis of breast tumors and their relationship with leukocyte characters and serum albumin level. Methods: A total of 75 serum samples and clinicopathological data of breast cancer patients from Tianjin Medical University Cancer Institute and Hospital were collected. The expression levels of 12 cytokines (IL-1β, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-17, TNF-α, IFN-α, IFN-γ) in serum were detected by immunofluorescence method. The absolute value of different kinds of leukocyte were determined by automatic blood analyzer. The concentration of albumin (ALB) was determined by bromocresol green method (BCG method). Univariate Cox regression and multivariate Cox regression were used to analyze the influence of the above indexes and clinicopathological features on the prognosis of patients. The correlation between the expression of cytokines and the absolute value of various kinds of leukocyte and serum albumin was studied by using multiple regression analysis. Results: A multivariate Cox proportional risk model was established by including breast cancer stage, molecular typing, IL-10, IL-12p70, absolute value of neutrophil, serum albumin and dipeptide aminopeptidase. It showed that different breast cancer stage (HR=0.415, 95%CI 0.191-0.902, P=0.026), and IL-10 (HR=0.177, 95%CI 0.038-0.821, P=0.027) were independent prognostic factors, Luminal B breast cancer had a statistically significant impact on survival compared with triple negative breast cancer (HR=0.193, 95%CI 0.067-0.554, P=0.002). The effect of other variables on survival was not statistically significant (P>0.05). Multiple linear regression analysis model demonstrated that the absolute value of neutrophils (b=1.498, t =2.938, P=0.005) and serum albumin (b=1.253, t=2.777, P=0.007) had statistically significant differences on IL-10, but breast cancer stage (b=0.635, t=1.776, P=0.08) and dipeptide aminopeptidase had no significant difference (b=0.952, t=1.715, P=0.091). Conclusions: Serum IL-10 plays an important role in indicating the prognosis of patients with breast cancer and its expression level was closely correlated with absolute value of neutrophil and serum albumin.

TRIM29 is differentially expressed in colorectal cancers of different primary locations and affects survival by regulating tumor immunity based on retrospective study and bioinformatics analysis.

In colorectal cancer (CRC) patients, different primary tumor locations caused distinct prognosis and clinicopathological features. It is necessary to identify specific tumor markers according tumor site. Our previous work has identified differentially expressed genes between CRC and adjacent normal tissues, in which only TRIM29 was differently expressed between right colon cancer (RCC) and left colon cancer (LCC) patients. Rectal cancer (RECC) was not included in this latter study and the effects of TRIM29 on the survival with RCC and LCC patients were not investigated. This study further verified TRIM29 expression through Gene Expression Omnibus (GEO) database and our retrospective study. The role of TRIM29 on survival according tumor sites was also explored. Furthermore, the molecular mechanisms of TRIM29 were explored. The GEO dataset was used to confirm the differential expression of TRIM29 in proximal and distal cancers. Moreover, TRIM29 were assess using immunohistochemistry (IHC) in 227 cases to observe the correlation between TRIM29 and tumor site. The relationship between TRIM29 and the clinicopathologic features was investigated according tumor sites. Furthermore, the disease-free survival (DFS) and overall survival (OS) was analyzed using the Kaplan-Meier method to assess the prognostic value of TRIM29. Finally, bioinformatics analysis was used to explore the molecular mechanisms. The Tumor-Immune System Interactions and Drug Bank database (TISIDB) was used to analyze the correlations between TRIM29 expression and tumor immune functions. The correlation of TRIM29 with tumor infiltrating lymphocytes or mismatch-repair-proficient/mismatch-repair-deficient (pMMR/dMMR) status was also investigated. TRIM29 expression was significantly higher in patients with RCC (P<0.001). RCC patients with high TRIM29 tended to be older, male, in stage III-IV, with N+ staging, and intestinal obstruction (P<0.001, P<0.001, P<0.001, P<0.001, and P=0.010, respectively). High TRIM29 expression was associated with an increased risk of recurrence/metastasis and death, only in RCC patients (P=0.020 and P<0.001). Functional annotations and immune activity analysis showed that TRIM29 is related to tumor infiltrating lymphocytes and immune dysfunction. TRIM29 plays varying roles in patients with different tumor sites. TRIM29 is correlated with the clinicopathological features and prognosis in RCC patients. Indeed, TRIM29 may serve as a new biomarker for RCC patients.

Clinical significance and correlation of PD-L1, B7-H3, B7-H4, and TILs in pancreatic cancer

BackgroundB7 molecules play significant roles in regulating tumor immunity, but their expression patterns and immuno-biological correlations in pancreatic cancer (PaCa) have not been fully discussed.MethodsRNA-sequencing data of B7 molecules of PaCa samples in the Cancer Genome Atlas (TCGA) dataset was downloaded from the UCSC Xena to assess the expression, correlation, and mutation of the B7 family in PaCa. Next, two PaCa tissue microarrays (TMAs, Cat. HPanA150CS02 and HPanA120Su02) were obtained from Outdo BioTech (Shanghai, China). To detect the expression levels of PD-L1, B7-H3 and B7-H4, immunohistochemistry (IHC) staining was performed on these TMAs.ResultsMost B7 molecules, including B7–1, B7–2, PD-L1, B7-DC, B7-H2, and B7-H5 exhibited similar expression patterns, but B7-H3, B7-H4, B7-H6, and B7-H7 showed outlier expression patterns compared with other B7 molecules. Besides, B7 molecules were genetically stable and exhibited low alteration frequency. IHC staining indicated PD-L1, B7-H3, and B7-H4 were up-regulated in PaCa tissues and showed uncorrelated expression patterns. Furthermore, high expression of PD-L1 and B7-H3 indicated poor-differentiated grades in PaCa. PD-L1 was positively, but B7-H4 was negatively correlated with CD8+ TILs infiltration in PaCa. Moreover, combined PD-L1 and B7-H4 expression was a novel subtyping strategy in PaCa, namely patients with both high PD-L1 and B7-H4 expression exhibited decreased CD8+ TILs infiltration in tumor tissues.ConclusionOverall, we systemically analyzed the expression patterns of B7 molecules and proposed a novel subtyping strategy in PaCa. Patients with both high PD-L1 and B7-H4 expression exhibited the immuno-cold phenotype, which may be not suitable for immunotherapy.

Recurrence- and Malignant Progression-Associated Biomarkers in Low-Grade Gliomas and Their Roles in Immunotherapy.

Despite a generally better prognosis than high-grade glioma (HGG), recurrence and malignant progression are the main causes for the poor prognosis and difficulties in the treatment of low-grade glioma (LGG). It is of great importance to learn about the risk factors and underlying mechanisms of LGG recurrence and progression. In this study, the transcriptome characteristics of four groups, namely, normal brain tissue and recurrent LGG (rLGG), normal brain tissue and secondary glioblastoma (sGBM), primary LGG (pLGG) and rLGG, and pLGG and sGBM, were compared using Chinese Glioma Genome Atlas (CGGA) and Genotype-Tissue Expression Project (GTEx) databases. In this study, 296 downregulated and 396 upregulated differentially expressed genes (DEGs) with high consensus were screened out. Univariate Cox regression analysis of data from The Cancer Genome Atlas (TCGA) yielded 86 prognostically relevant DEGs; a prognostic prediction model based on five key genes (HOXA1, KIF18A, FAM133A, HGF, and MN1) was established using the least absolute shrinkage and selection operator (LASSO) regression dimensionality reduction and multivariate Cox regression analysis. LGG was divided into high- and low-risk groups using this prediction model. Gene Set Enrichment Analysis (GSEA) revealed that signaling pathway differences in the high- and low-risk groups were mainly seen in tumor immune regulation and DNA damage-related cell cycle checkpoints. Furthermore, the infiltration of immune cells in the high- and low-risk groups was analyzed, which indicated a stronger infiltration of immune cells in the high-risk group than that in the low-risk group, suggesting that an immune microenvironment more conducive to tumor growth emerged due to the interaction between tumor and immune cells. The tumor mutational burden and tumor methylation burden in the high- and low-risk groups were also analyzed, which indicated higher gene mutation burden and lower DNA methylation level in the high-risk group, suggesting that with the accumulation of genomic mutations and epigenetic changes, tumor cells continued to evolve and led to the progression of LGG to HGG. Finally, the value of potential therapeutic targets for the five key genes was analyzed, and findings demonstrated that KIF18A was the gene most likely to be a potential therapeutic target. In conclusion, the prediction model based on these five key genes can better identify the high- and low-risk groups of LGG and lay a solid foundation for evaluating the risk of LGG recurrence and malignant progression.

Comprehensive Analysis of the Relationship between m6A Methylation Patterns and Immune Microenvironment in Lung Adenocarcinoma

背景与目的m6A RNA甲基化修饰在肺癌的发生与进展中起着重要作用，可以调节肿瘤免疫进而影响疾病预后。目前很多集中在某些特定的m6A效应器的差异表达及对肿瘤免疫细胞浸润的影响，但单个效应器表达的变化远远不足以反映m6A修饰特征的全貌，且关于m6A修饰对肺腺癌免疫微环境影响的研究仍较少。本研究拟探讨不同m6A修饰模式对肺腺癌中免疫微环境的影响。方法从癌症基因组图谱数据库（The Cancer Genome Atlas, TCGA）、加州大学圣克鲁兹分校泛癌全基因数据分析工具数据库（University of California Santa Cruz Xena, UCSC Xena）、基因表达综合数据库（Gene Expression Omnibus, GEO）获取肺腺癌相关数据信息。使用Maftools R分析肺腺癌队列中24个m6A效应器的基因突变，比较肺腺癌组织和正常组织中m6A效应器的表达差异，并通过Cox回归分析进行生存分析。通过Consensus Cluster Plus R非监督聚类的方法构建m6A修饰模式，进行m6A聚类生存分析、GSVA通路富集分析、免疫评分及免疫细胞浸润分析。在68例肺腺癌组织中，通过免疫组化分析LRPPRC蛋白表达水平与CD8、CD68的表达水平，验证LRPPRC与CD8+细胞毒性T细胞及巨噬细胞浸润的关系。结果在567例肺腺癌样本中有150例发生了m6A效应器突变，频率为26.46%。与正常组织相比，肺腺癌组织中共有6个读取器和3个写入器的表达明显上调。IGF2BP1和HNRNPC是影响肺腺癌患者预后的独立危险因素，且各效应器之间也存在大量的相互作用。构建了3种具有不同免疫细胞浸润特征和临床预后的m6A修饰模式。发现LRPPRC的表达与包括杀伤性T细胞和巨噬细胞在内的多种免疫细胞的浸润呈负相关，并在68例肺腺癌组织中得到验证。结论m6A修饰对肺腺癌免疫微环境的调节起着重要作用，LRPPRC有可能作为预测抗PD1免疫治疗效果的潜在生物标记物。

Comprehensive Analysis of the Immune Cell Infiltration Landscape and Immune-Related Methylation in Retinoblastoma.

Retinoblastoma is a common pediatric intraocular cancer, originating from cone precursors. The development of immunotherapies can help eradicate the tumor without vision loss, which would largely improve the quality of life of patients with retinoblastoma. Investigation of the tumor immune microenvironment provides knowledge for developing novel immunotherapies in cancer. However, the immune cell infiltrative landscape of retinoblastoma is unknown. Here, we compared the relative expression of immune gene signatures among 59 patients with retinoblastoma. The patients were divided into two subgroups according to the 28 types of immune cell infiltration (ICI) scores. We found that a subgroup with high ICI scores had increased expression levels of late cone markers, while the other subgroup exhibited larger tumor size and metastasis propensity. Furthermore, hypermethylated genes in the high-ICI subgroup were associated with immune regulation in the tumor microenvironment, suggesting that DNA methylation may play a vital regulatory role in retinoblastoma immunity. Our study provides a comprehensive framework for the systemic analysis of the influences of epigenetic events on the tumor immune microenvironment. We anticipate that our assay can not only provide insights into tumor immune regulation but also open up the perspectives for the identification of novel immunotherapy targets for retinoblastoma.

Transcriptional Patterns of Lower-Grade Glioma Patients with Distinct Ferroptosis Levels, Immunotherapy Response, and Temozolomide Sensitivity.

Background Many studies have defined a critical role for ferroptosis in cancer progression and therapy, but it is unclear how ferroptosis regulates tumor immunity or tumor microenvironment (TME). Methods In this study, 24 ferroptosis-regulators were assessed by nonnegative matrix factorization (NMF) consensus clustering to identify ferroptosis patterns in lower-grade gliomas (LGGs). Cell-type Identification By Estimating Relative Subsets Of RNA Transcripts (CIBERSORT) method and single sample gene set enrichment analysis (ssGSEA) were used to quantify immune cell infiltrations. The PCA algorithm was used to develop the ferroptosis-related score (FRscore) to measure ferroptosis levels. Results Two LGG subgroups named ferroptosis-related clusters 1 (FRC1) and 2 (FRC2), with distinct ferroptosis levels, immune infiltrations, and clinical outcomes were determined in 1,407 LGG samples. A well-designed scoring system was developed to evaluate the ferroptosis levels in LGG patients based on the FRSig gene profile and divided patients into low- and high-FRscore subgroups. Patients with low FRscores had lower ferroptosis levels and prolonged survival time and were expected to benefit from immune checkpoint blockade (ICB) therapy and showed higher sensitivity to TMZ chemotherapy. Findings also showed that the PI3K-AKT-mTOR pathway is activated by ferroptosis induction in SW1088 cells. Conclusions This study highlights the critical role of ferroptosis in TME formation and shaping, and quantitatively assessing ferroptosis levels in individual tumors can help to define the intratumor microenvironment and formulate precise treatment strategies for LGG patients.

CD4+ T cells mediate non-canonical rejection of major histocompatibility class-I deficient pancreatic tumors independently of CD8+ T cells

Abstract Cytolytic CD8+ T cells are a major mediator of immune-induced tumor rejection, but many patients have primary or acquired resistance to CD8+ T cell immunity. Reduced major histocompatibility complex I (MHC I) expression contributes to resistance, including treatment-refractory pancreatic ductal adenocarcinoma (PDAC). Interrogating interventions in PDAC will reveal novel mechanisms regulating sensitivity to immunotherapy. Using a genetically engineered mouse model of PDAC, we previously reported that agonistic anti-CD40 and dual immune checkpoint blockade (ICB) induces CD8+ and a CD4+ T cell responses mediating tumor rejection. Here, we utilized CRISPR− Cas9 to disrupt the dominant MHC I (H-2Kb) allele expression in PDAC cell lines, precluding direct antigen presentation to CD8+ T cells. Injection of MHC I-deficient tumor clones revealed normal tumor growth at baseline, and significantly delayed tumor growth in response to CD40/ICB in a CD4+ T cell-dependent manner (p &amp;lt; 0.0001 vs. vehicle-treated mice). There was no direct contribution of tumor rejection by CD8+ T cells, despite the generation of a tumor-specific response (24.5% vs. 1.98% in vehicle-treated mice, p&amp;lt;0.003). CD4+ T cells upregulated IFN-γ production (52.3% vs. 11.9% in vehicle-treated mice, p&amp;lt;0.0001), and host IFN-γ expression was required. No changes were observed in the expression of cytotoxic molecules by CD4+ T cells after CD40/ICB, and depletion of myeloid cells did not dampen treatment efficacy. Thus, CD4+ T cells mediated PDAC rejection via non-canonical mechanisms after treatment with CD40/ICB. This study provides critical insight into novel mediators of tumor clearance, and opportunities to exploit CD4 T+ cells as regulators of tumor immunity. Funded by the Parker Institute for Cancer Immunotherapy.

SLC7A11, a Potential Therapeutic Target Through Induced Ferroptosis in Colon Adenocarcinoma.

Background: Ferroptosis induced by SLC7A11 has an important translational value in the treatment of cancers. However, the mechanism of SLC7A11 in the pathogenesis of colon adenocarcinoma (COAD) is rarely studied in detail. Methods: SLC7A11 expression was explored with The Cancer Genome Atlas (TCGA), Gene Expression Omnibus (GEO) databases, and Western blot assay. The correlation of SLC7A11 expression with the abundance of infiltrating immune cells was evaluated via the TIMER database. The relation of SLC7A11 expression with immune cell markers was investigated via Gene Expression Profiling Interactive Analysis (GEPIA). The co-expression genes of SLC7A11 were screened by R packages, and the PPI was constructed via the STRING database. SLC7A11 and co-expressed gene modulators were selected by NetworkAnalyst and DSigDB database. The correlations between SLC7A11 and cancer immune characteristics were analyzed via the TIMER and TISIDB databases. Results: SLC7A11 is overexpressed in most tumors, including COAD. The expression level of SLC7A11 has a significant correlation with the infiltration levels of CD8+ T cells, neutrophils, and dendritic cells in COAD. The infiltrated lymphocyte markers of Th1 cell such as TBX21, IL12RB2, IL27RA, STAT1, and IFN-γ were strongly correlated with SLC7A11 expression. Five hub genes co-expressed with SLC7A11 that induce ferroptosis were identified, and mir-335-5p, RELA, and securinine have regulatory effects on it. SLC7A11 was negatively correlated with the expression of chemokines and chemokine receptors, such as CCL17, CCL19, CCL22, CCL23, CXCL14, CCR10, CX3CR1, and CXCR3, in COAD. Conclusion: SLC7A11 may play a role in induced ferroptosis and regulating tumor immunity, which can be considered as potential therapeutic targets in COAD.

The Role of Type-2 Conventional Dendritic Cells in the Regulation of Tumor Immunity.

Simple SummaryRecent studies revealed that type-2 conventional dendritic cells (cDC2s) play an important role in antitumor immunity by promoting cytotoxic T-cell responses and helper T-cell differentiation. This review outlines the role of cDC2s in tumor immunity and summarizes the latest progress regarding their potential in cancer vaccination and cDC2-targeted cancer immunotherapy.Conventional dendritic cells (cDCs) orchestrate immune responses to cancer and comprise two major subsets: type-1 cDCs (cDC1s) and type-2 cDCs (cDC2s). Compared with cDC1s, which are dedicated to the activation of CD8+ T cells, cDC2s are ontogenically and functionally heterogeneous, with their main function being the presentation of exogenous antigens to CD4+ T cells for the initiation of T helper cell differentiation. cDC1s play an important role in tumor-specific immune responses through cross-presentation of tumor-derived antigens for the priming of CD8+ T cells, whereas little is known of the role of cDC2s in tumor immunity. Recent studies have indicated that human cDC2s can be divided into at least two subsets and have implicated these cells in both anti- and pro-tumoral immune responses. Furthermore, the efficacy of cDC2-based vaccines as well as cDC2-targeted therapeutics has been demonstrated in both mouse models and human patients. Here we summarize current knowledge about the role of cDC2s in tumor immunity and address whether these cells are beneficial in the context of antitumor immune responses.

Cyclin-Dependent Kinase 6 Identified as the Target Protein in the Antitumor Activity of Tetrastigma hemsleyanum.

Background Tetrastigma hemsleyanum (T. hemsleyanum) is widely used as an adjuvant drug for tumor therapy but its antitumor therapeutic targets and molecular mechanisms have remained unclear. The prediction and analysis of natural products has previously used only network pharmacology methods to identify potential target proteins from public databases. In this study, we use comprehensive bioinformatics analysis and experimental verification to determine the antitumor mechanism of T. hemsleyanum.MethodsNetwork pharmacology analysis was used to predict the potential in vivo target proteins of T. hemsleyanum. The expression matrix and clinical data to perform an analysis of hub genes were collected from the TCGA and GTEx databases, specifically the analysis of expression, prognosis, tumor immune cell infiltration analysis, immune checkpoint genes, microsatellite instability, tumor mutational burden, tumor neoantigen, and immune microenvironment, which identify the roles and biological functions of the hub genes in pan-cancer. Finally, gene set enrichment analysis was used to verify the biological processes and signaling pathways involved in the pan-cancer expression profile.ResultsWe found 124 potential in vivo target proteins of T. hemsleyanum through network pharmacological analysis, and five hub genes (AKR1C1, MET, PTK2, PIK3R1, and CDK6) were then screened by protein–protein interaction (PPI) network analysis and molecular complex detection analysis (MCODE). Experimental intervention with an aqueous extract of T. hemsleyanum verified that these hub genes are the target proteins involved in the regulation of T. hemsleyanum in cells. A pan-cancer analysis then confirmed that CDK6 and MET are potential targets upon which T. hemsleyanum may exert antitumor action, especially in ACC, CESC, LGG, and PAAD. The CDK6 protein targeted by T. hemsleyanum is also involved in the immune and mutation process of pan-cancer, especially in the regulation of immune cell infiltration, immune checkpoint gene expression, microsatellite instability, tumor mutation burdens, and tumor neoantigens. Together, these analyses show that T. hemsleyanum affects tumor immune regulation and genomic stability. Finally, a gene set enrichment analysis confirmed that T. hemsleyanum regulates the cell cycle checkpoint.ConclusionsWe found that T. hemsleyanum can behave as an antitumor agent by acting as a potential cell cycle checkpoint inhibitor in CDK6-driven tumors, such as ACC, CESC, LGG, and PAAD, and that it acts as a tyrosine kinase receptor inhibitor that inhibits the expression of the proto-oncogene MET. Combined with an analysis of immune and mutation correlations in pan-cancer, we determined that T. hemsleyanum may function biologically as an immune regulator and interfere with the stability of the tumor genome, which is worthy of further study.

N6-methyladenosine (m6A) RNA modification in tumor immunity.

Growing evidence supports that cancer progression is closely associated with the tumor microenvironment and immune evasion. Importantly, recent studies have revealed the crucial roles of epigenetic regulators in shaping the tumor microenvironment and restoring immune recognition. N6-methyladenosine (m6A) modification, the most prevalent epigenetic modification of mammalian mRNAs, has essential functions in regulating the processing and metabolism of its targeted RNAs, and therefore affects various biological processes including tumorigenesis and progression. Recent studies have demonstrated the critical functions and molecular mechanisms underlying abnormal m6A modification in the regulation of tumor immunity. In this review, we summarize recent research progress in the potential roles of m6A modification in tumor immunoregulation, with a special focus on the anti-tumor processes of immune cells and involvement in immune-associated molecules and pathways. Furthermore, we review current knowledge regarding the close correlation between m6A-related risk signatures and the tumor immune microenvironment landscape, and we discuss the prognostic value and therapeutic efficacy of m6A regulators in a variety of cancer types.

Comprehensive analysis of N6-methyladenosine regulators with the tumor immune landscape and correlation between the insulin-like growth factor 2 mRNA-binding protein 3 and programmed death ligand 1 in bladder cancer

BackgroundN6-methyladenosine (m6A) is one of the most abundant post-transcriptional modifications of RNA. However, there is limited information about the potential roles of m6A regulators in tumor immunity. Therefore, in this study, we aimed to testify the functions of m6A regulators in bladder cancer as well as their association with the tumor immune landscape.MethodsWe reported the variation and expression levels of m6A regulators in the TCGA database and GTEx database of bladder cancer. Clusters, risk score patterns, and nomograms were constructed to evaluate the function and prognostic value of m6A regulators. Furthermore, we constructed nomogram to evaluate the prognosis of the individual patients. The correlation between insulin-like growth factor 2 mRNA-binding protein 3 (IGF2BP3) and programmed cell death ligand 1 (PD-L1) was evaluated both in vitro and in vivo.ResultsWe found that the tumor grade and DNA damage pathways were strongly correlated with distinct clusters. Furthermore, two risk score groups with six m6A regulators were identified using the least absolute shrinkage and selection operator (LASSO) and multivariable Cox regression analysis, which could be regarded as independent prognostic markers in patients with bladder cancer. The risk score pattern was linked to the tumor immune landscape, indicating a correlation between immune checkpoints and m6A regulators. Moreover, an m6A regulator, IGF2BP3, was found to be highly expressed in the tumor samples, regulating both the total and membrane-bound PD-L1 expression levels.ConclusionsThe results of this study revealed that the m6A clusters and patterns play crucial roles in the regulation of tumor immunity, which may be used to develop comprehensive treatment strategies for the management of bladder cancer.

FCER1G positively relates to macrophage infiltration in clear cell renal cell carcinoma and contributes to unfavorable prognosis by regulating tumor immunity

BackgroundTumor-associated macrophages (TAMs) are closely related to unfavorable prognosis of patients with clear cell renal cell carcinoma (ccRCC). However, the important molecules in the interaction between ccRCC and TAMs are unclear.MethodsTCGA-KIRC gene expression data of tumor tissues and normal tissues adjacent to tumor were compared to identify differentially expressed genes in ccRCC. TAMs related genes were discovered by analyzing the correlation between these differentially expressed genes and common macrophage biomarkers. Gene set enrichment analysis was performed to predict functions of TAMs related gene. The findings were further validated using RNA sequencing data obtained from the CheckMate 025 study and immunohistochemical analysis of samples from 350 patients with ccRCC. Kaplan–Meier survival curve, Cox regression analysis and Harrell’s concordance index analysis were used to determine the prognostic significance.ResultsIn this study, we applied bioinformatic analysis to explore TAMs related differentially expressed genes in ccRCC and identified 5 genes strongly correlated with all selected macrophage biomarkers: STAC3, LGALS9, TREM2, FCER1G, and PILRA. Among them, FCER1G was abundantly expressed in tumor tissues and showed prognostic importance in patients with ccRCC who received treatment with Nivolumab; however, it did not exhibit prognostic value in those treated with Everolimus. We also discovered that high expression levels of FCER1G are related to T cell suppression. Moreover, combination of FCER1G and macrophage biomarker CD68 can improve the prognostic stratification of patients with ccRCC from TCGA-KIRC. Based on the immunohistochemical analysis of samples from patients with ccRCC, we further validated that FCER1G and CD68 are both highly expressed in tumor tissue and correlate with each other. Higher expression of CD68 or FCER1G in ccRCC tissue indicates shorter overall survival and progression-free survival; patients with high expression of both CD68 and FCER1G have the worst outcome. Combining CD68 and FCER1G facilitates the screening of patients with a worse prognosis from the same TNM stage group.ConclusionsHigh expression of FCER1G in ccRCC is closely related to TAMs infiltration and suppression of T cell activation and proliferation. Combining the expression levels of FCER1G and macrophage biomarker CD68 may be a promising postoperative prognostic index for patients with ccRCC.

Roles of the Exosomes Derived From Myeloid-Derived Suppressor Cells in Tumor Immunity and Cancer Progression.

Tumor immunity is involved in malignant tumor progression. Myeloid-derived suppressor cells (MDSCs) play an irreplaceable role in tumor immunity. MDSCs are composed of immature myeloid cells and exhibit obvious immunomodulatory functions. Exosomes released by MDSCs (MDSCs-Exos) have similar effects to parental MDSCs in regulating tumor immunity. In this review, we provided a comprehensive description of the characteristics, functions and mechanisms of exosomes. We analyzed the immunosuppressive, angiogenesis and metastatic effects of MDSCs-Exos in different tumors through multiple perspectives. Immunotherapy targeting MDSCs-Exos has demonstrated great potential in cancers and non-cancerous diseases.

BTBD10 is a Prognostic Biomarker Correlated With Immune Infiltration in Hepatocellular Carcinoma.

Background: BTBD10 serves as an activator of Akt family members through decreasing the protein phosphatase 2A-mediated dephosphorylation. The present study attempted to investigate the prognostic value of BTBD10 in hepatocellular carcinoma (HCC), specially, its relationship with tumor-infiltrating lymphocytes (TILs). Methods: BTBD10 expression was evaluated in HCC using The Cancer Genome Atlas (TCGA) and Xijing Hospital database, and verified in HCC cell lines. Cox analyses were performed to analyze independent prognostic risk factors for HCC. The optimal cut-off value of BTBD10 was calculated, by which all patients were divided into two groups to compare the overall survival (OS). The signaling pathways were predicted, by which BTBD10 may affect the progression of HCC. To investigate the impact of BTBD10 on HCC immunotherapy, correlations between BTBD10 and TILs, immune checkpoints, m6A methylation-related genes and ferroptosis-related genes were assessed. The distribution of half-maximal inhibitory concentration (IC50) of diverse targeted drugs was observed based on the differential expression of BTBD10. Results: BTBD10 expression was higher in HCC tissues and cell lines than that of normal liver tissues and cells. The patients with high expression of BTBD10 showed a worse OS, as compared to that of BTBD10 low-expressing group. Cox analyses indicated that BTBD10 was an independent prognostic risk factor for HCC. Several molecular pathways of immune responses were activated in HCC patients with high-expressing of BTBD10. Furthermore, BTBD10 expression was demonstrated to be positively correlated with tumor-infiltrating B cells, T cells, macrophages, neutrophils and dendritic cells. Meanwhile, the expression of BTBD10 was synchronized with that of several m6A methylation-related genes, ferroptosis-related genes and immune checkpoints. The IC50 scores of Sorafenib, Navitoclax, Veliparib, Luminespib, and Imatinib were found to be lower in BTBD10 high-expressing HCC group. Conclusion: BTBD10 negatively regulates tumor immunity in HCC and exhibits adverse effect on the prognosis of HCC, which could be a potential target for immunotherapy.