Immune Response In Tumor Microenvironment Research Articles

Abstract PO4-28-05: Prolactin drives labile iron transfer from macrophages to breast cancer cells through CD44

Abstract Cancer cells are “ferrophilic” and exhibit great dependence on iron. While iron metabolism is closely related to tumor development, the source of iron and the mechanisms cancer cells adopt to actively acquire iron is not well understood. In the present study, we discovered a novel function of the lactating hormone, prolactin, in the regulation of iron transport. In breast cancer cells using mouse triple negative breast cancer cell lines EO771 and Py230 for demonstration, prolactin stimulation increased intracellular labile iron pool. Analyses of gene expression involved in iron transport revealed dramatic induction of CD44, a surface receptor for iron bound hyaluronan. Other genes involved in iron uptake including transferrin receptor (TFRC), CD163 and divalent metal transporter 1 (DMT1) were not changed. The utilization of neutralizing antibodies against CD44 significantly blocked prolactin mediated labile iron pool accumulation in breast cancer cells. In contrast to what was found in breast cancer cells, prolactin had the opposite impact on immune macrophages. Prolactin treatment increased ferroportin (FPN I, iron exporter) but had no influence on all iron uptake transporters leading to net iron release by macrophages. To further determine if the iron released by macrophages can be a direct source of iron utilized by breast cancer cells, we conducted co-culture work of macrophages with pre-stained labile iron pool and breast cancer cells without any iron staining and found a gradual increase of fluorescent iron pool developed in breast cancer cells over time. In conclusion, our work presents a multidisciplinary regulation of iron transport among prolactin, macrophages, and cancer cells. The novel regulatory role of prolactin to drive iron flow can provide new information on fine-tuning immune responses in tumor microenvironment and potentially benefit the development of novel therapeutics. Citation Format: Reagan Farrell, Nicholas Pascuzzi, Yi-Ling Chen, Miguel Torres, Mary Kim, Lauren Gollahon, Kuan-Hui Chen. Prolactin drives labile iron transfer from macrophages to breast cancer cells through CD44 [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-28-05.

Association of Interleukin-12 and Interleukin-18 Polymorphisms with Acute Lymphoblastic Leukemia Disease in Iraqi Patients

Acute lymphoblastic leukemia (ALL) is a blood cancer where high numbers of abnormal, immature lymphocytes called blasts start over-multiplying in the bone marrow. The lymphocyte cells and interleukins are crucial for controlling the immune response in tumor microenvironment. Many of the single nucleotide polymorphisms (SNPs) of some interleukin genes may change protein synthesis or function and regulate the immune response. A lot of these changes have been connected to a high risk of developing cancer. Aim of this study was to investigate the potential association between IL-12 (+1188 A/C) (rs3212227) and IL-18 (–607 A/C) (rs1946518) polymorphism with serum levels of interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α) and their genotypes association with the susceptibility to ALL disease in Iraqi patients. The study included 59 ALL patients and 30 healthy controls. The detection of IL-12 and IL-18 SNP genes was determined by ARMS-PCR methods and serum levels of IFN-γ and TNF-α were determined by utilizing enzyme linked immuno-sorbent assay (ELISA) kits. Results observed no significant association of IL-12 and IL-18 polymorphism with serum IFN-γ and TNF-α level. Genotyping frequencies of IL-12 (+1188A/C) CA, AA and CA+AA genotypes showed a strong association with the development of ALL disease, IL-18 genotypes showed no significant association with the disease progression.

The Regulation of FOXP3 Expression and Its Therapeutic Potential in Non-Small Lung Cancer

Non-small cell cancer (NSCLC) is today one of the most threatening malignant tumors, and it is still rarely cured by current standard therapies with a high mortality rate. The transcription factor Forkhead box p3 (FOXP3) is a potential molecular target for NSCLC since it has been identified with an oncogenic effect in the tumor microenvironment (TME). It is well known that FOXP3 expression could characterize the regulatory T cells (Treg), which plays a critical role in regulating anti-tumor immune responses in TME. Through analyzing the basic mechanisms of FOXP3 and Treg in distinct tumor tissues, they have been found with contradictory expression and the frequency in different TME. In NSCLC, the suppressive function of Treg to immune systems and frequency of FOXP3 is accumulated to promote the development of tumors. Most recently, accumulating evidence reveals the therapeutic potential of targeting FOXP3 and the related signaling pathways in the treatment of NSCLC patients. Although unsolved side effects and limitations that still must be considered when targeting FOXP3 and FOXP3-related proteins, there is no doubt that they are potential therapeutic applications for NSCLC treatment. The main objective of this review is to mention the structure, function and recent progress in treatment targeting FOXP3 and FOXP3-related signaling pathways, especially to therapies with immune checkpoint inhibitors.

Genes associated with cellular senescence favor melanoma prognosis by stimulating immune responses in tumor microenvironment

PurposeSkin cutaneous melanoma (SKCM), a malignant tumor from melanocytes, is the fifth most prevalent tumor. Immune checkpoint inhibitor (ICI) immunotherapy improves prognosis of SKCM, but immune response varies for different populations. Cellular senescence in the tumor microenvironment (TME) promotes antitumor immunity, mediated by dendritic cells (DC) and CD8+ T cells. Therefore, we sought to explore the role of cellular senescence in the TME of SKCM through bioinformatics and machine learning. MethodsFirst, we obtained 93 cellular senescence-prognosis genes (CSPGs) by univariate survival analysis. Thereafter, 23 optimal CSPGs were obtained by least absolute shrinkage and selection operator (lasso) analysis. Based on the riskscore obtained by lasso analysis and clinical information from multivariate cox, we obtained the nomogram of SKCM, which was validated in the validation cohort. Based on the riskscore, the patients were split into low- and high-risk groups. Functional differences between the two groups were analyzed using Metascape and GSEA, and immune infiltration differences were achieved by multiple algorithms. We obtained a risk prediction nomogram for the validated SKCM based on the lasso model by univariate and multivariate cox regression analysis. ResultsIn the low-risk group, immune responses were in an active state. NK, CD8+ T, DC, macrophages, and neutrophils were significantly upregulated, and ICI‐relevant genes were notably upregulated. With the differentially expressed genes (DEGs) and optimal CSPGs, we obtained the hub genes: NOX4, NTN4, PROX1, and TRPM8. The hub genes were mainly expressed by cancer-associated fibroblasts (CAFs) and endothelial cells by single cell analysis, which were mainly associated with angiogenesis. ConclusionGenes associated with cellular senescence favor SKCM prognosis by stimulating immune responses in TME. Patients with high expression of cellular senescence associated genes in the TME might have better benefit from ICI immunotherapy. Cellular senescence functions as a pro-tumor agent in mesenchymal cells and needs further study.

Abstract 1546: ADCC enhanced anti-CD25 mAb (TST010) demonstrated antitumor activity via depleting Treg cells and increasing CD8+T/Treg ratio in preclinical tumor models

Abstract Regulatory T (Treg) cells are a distinctive lineage of CD4+ T cells as characterized by expression of FoxP3 and CD25 markers. Treg cells are involved in tumor development and progression by inhibiting antitumor immunity. Accumulation of Treg cells in the tumor microenvironment (TME) is often associated with an adverse prognosis in multiple cancers and also has implication for predicting response to immunotherapy. Treg cell depletion strategy has been explored in preclinical and clinical settings to induce effective antitumor immune responses in TME. CD25, a high-affinity binding subunit alpha of the IL-2 receptor, is constitutively and highly expressed on Treg cells but also transiently upregulated on effector T cells. The high level of CD25 expression on Treg cells could deprive of IL-2 for effector T cells and inhibit their proliferation. Thus, an effective approach to selectively suppress Treg cell function is to deplete Treg cells while avoid blocking IL-2 binding to CD25 for allowing T effect cell proliferation and activation. Using our immune tolerance breaking antibody platform, we generated a humanized IgG1 subtype anti-CD25 monoclonal antibody (TST010). TST010 bound to human CD25 with high affinity and selectivity but did not block IL-2 binding to CD25. No blocking activity to IL-2 signaling on human PBMC was further confirmed by the phosphorylation of Stat5 using FACS analysis. By reducing fucosylation during cell culture process, TST010 mAb gained the enhanced ADCC activity. In an ADCC reporter assay, TST010 showed its sub-nanomolar ADCC activity against CD25-overexpression SU-DHL-1 cell line and human induced Treg cells, but much less to human activated CD8+ T effect cells due to lower CD25 expression. Similarly, in a NK cytotoxicity assay, TST010 selectively killed human induced Treg cells in contrast to activated CD8+ T cells. In vivo antitumor activity of TST010 was evaluated in the syngeneic mouse tumor models. The mouse colon cancer MC38 model was developed on the human CD25 knock-in mice. In the MC38 tumor model, TST010 markedly reduced the Treg (FoxP3+CD4+) population in both peripheral blood and TME. Importantly, the CD8+T/Treg ratio in tumor was increased starting at 2 weeks post TST010 treatment, and it resulted in a significant single agent tumor growth inhibition at a 10 mg/kg dose. Furthermore, in the LLC tumor model which does not respond well to checkpoint inhibitors, the anti-CD25 mAb showed a good combination effect with a PD-L1 inhibitor. In summary, we have discovered a novel therapeutic anti-CD25 mAb that can deplete Treg cells without blocking IL-2 signaling. As demonstrated in our preclinical tumor models, it has a good potential to induce effective antitumor immune responses in TME and tumor growth inhibition especially in combination with PD-1/PD-L1 treatment. Citation Format: Di Sun, Huanhuan Guo, Hongjun Li, Xinlai Yao, Yiqing Xu, Shenjie Zhang, Shijie Zhou, Xiaoli Zi, Lisa Zheng, Yi Gu, Xueming Qian. ADCC enhanced anti-CD25 mAb (TST010) demonstrated antitumor activity via depleting Treg cells and increasing CD8+T/Treg ratio in preclinical tumor models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1546.

Immune checkpoint inhibitors as mediators for immunosuppression by cancer-associated fibroblasts: A comprehensive review

The tumor microenvironment (TME) is a significant contributor to cancer progression containing complex connections between cellular and chemical components and provides a suitable substrate for tumor growth and development. Growing evidence shows targeting tumor cells while ignoring the surrounding TME is not effective enough to overcome the cancer disease. Fibroblasts are essential sentinels of the stroma that due to certain conditions in TME, such as oxidative stress and local hypoxia, become activated, and play the prominent role in the physical support of tumor cells and the enhancement of tumorigenesis. Activated fibroblasts in TME, defined as cancer-associated fibroblasts (CAFs), play a crucial role in regulating the biological behavior of tumors, such as tumor metastasis and drug resistance. CAFs are highly heterogeneous populations that have different origins and, in addition to their role in supporting stromal cells, have multiple immunosuppressive functions via a membrane and secretory patterns. The secretion of different cytokines/chemokines, interactions that mediate the recruitment of regulatory immune cells and the reprogramming of an immunosuppressive function in immature myeloid cells are just a few examples of how CAFs contribute to the immune escape of tumors through various direct and indirect mechanisms on specific immune cell populations. Moreover, CAFs directly abolish the role of cytotoxic lymphocytes. The activation and overexpression of inhibitory immune checkpoints (iICPs) or their ligands in TME compartments are one of the main regulatory mechanisms that inactivate tumor-infiltrating lymphocytes in cancer lesions. CAFs are also essential players in the induction or expression of iICPs and the suppression of immune response in TME. Based on available studies, CAF subsets could modulate immune cell function in TME through iICPs in two ways; direct expression of iICPs by activated CAFs and indirect induction by production soluble and then upregulation of iICPs in TME. With a focus on CAFs’ direct and indirect roles in the induction of iICPs in TME as well as their use in immunotherapy and diagnostics, we present the evolving understanding of the immunosuppressive mechanism of CAFs in TME in this review. Understanding the complete picture of CAFs will help develop new strategies to improve precision cancer therapy.

Role of hypoxia in the tumor microenvironment and targeted therapy.

Tumor microenvironment (TME), which is characterized by hypoxia, widely exists in solid tumors. As a current research hotspot in the TME, hypoxia is expected to become a key element to break through the bottleneck of tumor treatment. More and more research results show that a variety of biological behaviors of tumor cells are affected by many factors in TME which are closely related to hypoxia. In order to inhibiting the immune response in TME, hypoxia plays an important role in tumor cell metabolism and anti-apoptosis. Therefore, exploring the molecular mechanism of hypoxia mediated malignant tumor behavior and therapeutic targets is expected to provide new ideas for anti-tumor therapy. In this review, we discussed the effects of hypoxia on tumor behavior and its interaction with TME from the perspectives of immune cells, cell metabolism, oxidative stress and hypoxia inducible factor (HIF), and listed the therapeutic targets or signal pathways found so far. Finally, we summarize the current therapies targeting hypoxia, such as glycolysis inhibitors, anti-angiogenesis drugs, HIF inhibitors, hypoxia-activated prodrugs, and hyperbaric medicine.

The role of lipid metabolism in tumor immune microenvironment and potential therapeutic strategies.

Aberrant lipid metabolism is nonnegligible for tumor cells to adapt to the tumor microenvironment (TME). It plays a significant role in the amount and function of immune cells, including tumor-associated macrophages, T cells, dendritic cells and marrow-derived suppressor cells. It is well-known that the immune response in TME is suppressed and lipid metabolism is closely involved in this process. Immunotherapy, containing anti-PD1/PDL1 therapy and adoptive T cell therapy, is a crucial clinical cancer therapeutic strategy nowadays, but they display a low-sensibility in certain cancers. In this review, we mainly discussed the importance of lipid metabolism in the formation of immunosuppressive TME, and explored the effectiveness and sensitivity of immunotherapy treatment by regulating the lipid metabolism.

Abstract 5594: Systematic identification of the chemo-immunotherapy synergism to overcome immunotherapy resistance in cancer

Abstract Immune checkpoint blockades (ICB) have prompted a revolution in cancer treatment, but its low response rate and resistance remain a major problem. Clinical trials have reported the combination of ICB with chemotherapy can improve patient treatment outcome, but the synergy potential of most chemo drugs with ICBs are not systematically evaluated due to the limited insight of drug induced alterations in tumor microenvironment. Here, we reported a novel algorithm termed post-treatment-based synergy prediction for chemotherapy and ICB combination (POSEIDON). POSEIDON integrates one million drug-induced transcriptomic changes of T cell and cancer cells from the Library of Integrated Network-Based Cellular Signatures (LINCS) and ICB-induced single cell transcriptomic changes of 500K cells from tumor microenvironments across multiple cancer types. Using the gene expression signatures induced by chemotherapy and immunotherapy, POSEIDON utilized a reversal-orthogonal framework to predict the chemo compound that can potentially overcome ICB resistance and improve treatment beneficial. In an anti-PD1 treated melanoma patient cohort, POSEIDON identified a group of VEGFR inhibitors and HDAC inhibitors, including tivozanib and mocetinostat, showing the highest synergy potential with anti-PD1 therapy to reverse the anti-PD1 resistance through regulating T cell activities. In a triple negative breast cancer patient cohort who received the combination of anti-PDL1 and paclitaxel, POSEIDON also predicted several HDAC inhibitors (e.g., mocetinostat) and thymidylate synthase inhibitors (e.g., trifluridine) to synergize with the combination of anti-PDL1 plus paclitaxel and reverse the combination resistance through activating the immune response in tumor microenvironment. In addition to the synergy through tumor microenvironment, POSEIDON is also capable of predicting the synergy through tumor intrinsic mechanism. POSEIDON identified the genetic inhibition of EZH2 can potentially reverse anti-PD1 resistance via upregulation of tumor antigen presentation and interferon signaling. The synergy between EZH2 inhibitor and anti-PD1 was further validated in vivo by treating the anti-PD1 resistant 4T1.2 mouse model with FDA approved EZH2 inhibitor tazemetostat and PD1 antibody. Our data showed that, while tazemetostat or anti-PD1 alone failed to inhibit tumor growth, the combination of tazemetostat and anti-PD1 can significantly reduce the tumor size and increase overall survival. Collectively, our study demonstrated that POSEIDON can provide reliable prediction of chemo-immunotherapy synergism by integrating the post-treatment transcriptomic profiles from patient tumor microenvironment and cell lines. We envision that the systematic identification of chemo-immunotherapy synergism will help cancer patients to cope with immunotherapy resistance in the short future. Citation Format: Yue Wang, Yifei Wang, Xiaofei Wang, Zehua Wang, Min Zhang, Da Yang. Systematic identification of the chemo-immunotherapy synergism to overcome immunotherapy resistance in cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5594.

Lenvatinib combined with nivolumab in advanced hepatocellular carcinoma-real-world experience

Lenvatinib, a multi-tyrosine kinase inhibitor that inhibits vascular endothelial growth factor and fibroblast growth factor receptors pathway, activated the immune response in tumor microenvironment. However, the combination of lenvatinib and anti-PD-1 has been reported in early phase studies. Hence, this study aims to explore the efficacy and toxicity of lenvatinib combined with nivolumab in the real-world setting. Advanced HCC patients who underwent lenvatinib combined with nivolumab (L + N group) treatment at Taipei Veterans General Hospital (Taipei, Taiwan) were reviewed between January 2016 and December 2020. Treatment response and outcomes were collected and analyzed. A control group with lenvatinib (L group) was also included for comparison. Forty patients were included in L + N group and 47 in L group. The L + N group demonstrated a higher objective response rate than L group (45.0% vs. 23.4%, p = 0.03). The L + N group also achieved longer PFS (7.5 vs. 4.8 months, p = 0.05) and OS (22.9 vs. 10.3 months, p = 0.01) than L group. Patients with HBV infection and REFLECT criteria fit demonstrated a trend of better prognosis. The PFS for those with PR, SD and PD groups were 11.2, 6.4, and 2.2 months and OS were non-reached, 14.6 and 4.7 months, respectively. Portal vein thrombosis (HR 4.3, 95% C.I. 1.5–12.8) and AFP > 400 ng/mL (HR 3.3, 95% C.I. 1.1–9.3) were poor prognostic factors and nivolumab used remained a protective factor (HR 0.2, 95% C.I. 0.1–0.7). Dermatitis (35.0%), pruritis (27.5%), and hypothyroidism (27.5%) were the common toxicities. Few patients developed grade 3/4 toxicities, including dermatitis (15%), gastrointestinal bleeding (7.5%), hypertension (5.0%), pneumonitis (2.5%) and stomatitis (2.5%). This is the first real-world data reporting the promising efficacy and tolerable toxicities of lenvatinib combined with nivolumab in advanced HCC. Further randomized trials are prompted.

The Role of Beetroot Extract in Overcoming Chemoresistance of Neoadjuvant Adriamycin Cyclophosphamide Regimen by Targeting Immune Response in Tumor Microenvironment: A Preclinical Study in Mammary Adenocarcinoma Rats.

Objective:This study aims to determine the role of beetroot extract in overcoming the chemoresistance of Neoadjuvant Adriamycin Cyclophosphamide (NAC) regimens with a target immune response in the tumour microenvironment at the pre-clinical stage. Methods:This study was conducted on rats with 7,12-Dimethyl Benz (α) Anthracene (DMBA) induced mammary adenocarcinoma. Adriamycin Cyclophosphamide was given in 4 cycles, whereas beetroot extract was administered three times each cycle. Observations of CD8 T cells and Myeloid Derivative Suppressive Cells (MDSC) expression levels and pathological responses were carried out on tumour tissue taken at the end of the observation. Results:Supplementation of beetroot extract to NAC could significantly increase CD8 T cells and decrease MDSC in the tumour microenvironment. The addition of beetroot extract gave a better pathological response. Conclusion:Beetroot extract enhances the immune response in the tumor microenvironment so that it has the potential to overcome chemoresistance in NAC.

MTOR-Mediated Regulation of Immune Responses in Cancer and Tumor Microenvironment.

The mechanistic/mammalian target of rapamycin (mTOR) is a downstream mediator in the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathways, which plays a pivotal role in regulating numerous cellular functions including cell growth, proliferation, survival, and metabolism by integrating a variety of extracellular and intracellular signals in the tumor microenvironment (TME). Dysregulation of the mTOR pathway is frequently reported in many types of human tumors, and targeting the PI3K/Akt/mTOR signaling pathway has been considered an attractive potential therapeutic target in cancer. The PI3K/Akt/mTOR signaling transduction pathway is important not only in the development and progression of cancers but also for its critical regulatory role in the tumor microenvironment. Immunologically, mTOR is emerging as a key regulator of immune responses. The mTOR signaling pathway plays an essential regulatory role in the differentiation and function of both innate and adaptive immune cells. Considering the central role of mTOR in metabolic and translational reprogramming, it can affect tumor-associated immune cells to undergo phenotypic and functional reprogramming in TME. The mTOR-mediated inflammatory response can also promote the recruitment of immune cells to TME, resulting in exerting the anti-tumor functions or promoting cancer cell growth, progression, and metastasis. Thus, deregulated mTOR signaling in cancer can modulate the TME, thereby affecting the tumor immune microenvironment. Here, we review the current knowledge regarding the crucial role of the PI3K/Akt/mTOR pathway in controlling and shaping the immune responses in TME.

Abstract P4-04-04: APOBEC3F is expressed by immune cells and is associated with immune response and survival in triple negative breast cancer

Abstract Background: The Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) cytidine deaminases subfamily has been a focus of attention as a cause of characteristic somatic mutations in breast cancer. APOBEC3A and APOBEC3B are known to be the key mutation drivers in breast cancer, while the other APOBEC3 family members have been implicated in infection. APOBEC3F was reported to inhibit HIV replication, but its role in cancer has not been reported. We hypothesized that APOBEC3F is associated with infiltrating immune cells and immune responses in tumor microenvironment, and with survival of breast cancer patients. Method: We analyzed a total of 2978 breast cancer patients’ clinical data and transcriptome from the Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) and The Cancer Genome Atlas (TCGA) datasets. We also used a total of 21802 single-cell transcriptomes from the GSE75688 and GSE114725 single-cell sequencing datasets of the primary breast cancer cohort. We defined "high expression" as top two thirds of APOBEC3F expression in each cohort. Result: APOBEC3F expression was significantly high in triple negative (TNBC), which is known to have the most abundant immune cell infiltration among the subtypes. Disease-specific survival (DSS) and overall survival (OS) were consistently better in high APOBEC3F TNBC (all p < 0.05), but not in luminal subtype, that led us to hypothesize that APOBEC3F may enhance cancer immunity. APOBEC3F expression was not consistently associated with intratumor heterogeneity, mutation rates nor neoantigens that are all reflections of cancer cells in both TNBC and luminal breast cancer, except for HRD in TNBC (p = 0.024). APOBEC3F expression did not correlate with response to any of the drugs tested in breast cancer cell lines in vitro. On the other hand, APOBEC3F expression was consistently associated with multiple types of immune cell infiltrations, such as CD8+ T-cells, M1 and M2 macrophage, Dendritic cells, Natural Killer cells (NK), and B-cells (all p < 0.05) with high CYT (p < 0.001) regardless of subtype. In TNBC, high APOBEC3F tumor significantly enriched only immune-related gene sets; inflammatory response, IL-2/STAT5 signaling, IFN γ response, IFN α response,IL-6/JAC/STAT3 signaling and complement, consistently in both cohorts, and associated with high levels of immune activity scores such as IFN-γ response, tumor infiltrating lymphocytes regional fraction, lymphocyte infiltration signature, leukocyte fraction, and TCR richness in TCGA cohort (all p < 0.05). In luminal breast cancer, high APOBEC3F tumor significantly enriched not only immune-related gene sets similar to TNBC, but also tumor aggravating gene sets such as KRAS signaling, PI3K signaling, angiogenesis, metabolism-related gene sets, as well as apoptosis-related gene sets. We further found that APOBEC3F was exclusively expressed in immune cells compared to myeloid, stromal, and cancer cells using the GSE75688 breast cancer single cell sequencing cohort, and highly expressed in CD8, CD4, NK, and B-cells, and neutrophils in GSE114725 breast cancer single cell sequencing cohort (both p < 0.001). High APOBEC3F immune cells were significantly associated with higher expression of IFN-γ, cytolytic activity score and its component genes, and with immune cell proliferation (all p < 0.001). High APOBEC3F tumor was associated with significantly elevated levels of all the immune checkpoint molecules tested (all p < 0.001). Conclusion: Our study demonstrated that high APOBEC3F gene expression is exclusively expressed in immune cells and is associated with immune response, immune cytolytic activity in the tumor microenvironment, and with favorable survival in triple-negative breast cancer using in silico bioinformatic analyses of multiple independent large patient cohorts. Citation Format: Rongrong Wu, Masanori Oshi, Mariko Asaoka, Akimitsu Yamada, Li Yan, Itaru Endo, Takashi Ishikawa, Kazuaki Takabe. APOBEC3F is expressed by immune cells and is associated with immune response and survival in triple negative breast cancer [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P4-04-04.

Screening and Identification of Novel Potential Biomarkers for Breast Cancer Brain Metastases.

Brain metastases represent a major cause of mortality among patients with breast cancer, and few effective targeted treatment options are currently available. Development of new biomarkers and therapeutic targets for breast cancer brain metastases (BCBM) is therefore urgently needed. In this study, we compared the gene expression profiles of the brain metastatic cell line MDA-MB-231-BR (231-BR) and its parental MDA-MB-231, and identified a total of 84 genes in the primary screening through a series of bioinformatic analyses, including construction of protein-protein interaction (PPI) networks by STRING database, identification of hub genes by applying of MCODE and Cytohubba algorithms, identification of leading-edge subsets of Gene Set Enrichment Analysis (GSEA), and identification of most up-regulated genes. Eight genes were identified as candidate genes due to their elevated expression in brain metastatic 231-BR cells and prognostic values in patients with BCBM. Then we knocked down the eight individual candidate genes in 231-BR cells and evaluated their impact on cell migration through a wound-healing assay, and four of them (KRT19, FKBP10, GSK3B and SPANXB1) were finally identified as key genes. Furthermore, the expression of individual key genes showed a correlation with the infiltration of major immune cells in the brain tumor microenvironment (TME) as analyzed by Tumor Immune Estimation Resource (TIMER) and Gene Expression Profiling Interactive Analysis (GEPIA), suggesting possible roles of them in regulation of the tumor immune response in TME. Therefore, the present work may provide new potential biomarkers for BCBM. Additionally, using GSEA, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) Enrichment Analysis, we determined the top enriched cellular functions or pathways in 231-BR cells, which may help better understand the biology governing the development and progression of BCBM.

Resveratrol attenuates TNBC lung metastasis by down-regulating PD-1 expression on pulmonary T cells and converting macrophages to M1 phenotype in a murine tumor model

Triple-negative breast cancer (TNBC) is an invasive breast cancer with the characteristics of easy to develop distant metastasis. Immune escape is one of the main reasons for TNBC growth and metastasis. Enhancement of T cell-mediated anti-tumor activity may benefit to inhibit tumor metastasis and improve the efficacy of cancer therapy. As a natural bioactive substance, resveratrol shows potential capability to prevent or suppress the development of a variety of cancers through direct or indirect effects, including immunoregulatory effect. However, whether resveratrol might affect lung metastasis of TNBC, and whether the effect of resveratrol might be associated with resveratrol-regulated immune responses in tumor microenvironment is still unknown. In this study, by using an experimental metastatic mouse 4 T1 tumor model, we identified that resveratrol may suppress TNBC lung metastasis by elevating local anti-tumor immunity. Indeed, an increase in the cytotoxic activity of CD8+T cells as well as the levels of type 1 cytokine IFN-γ and IL-2 in the lungs of resveratrol-treated tumor bearing mice were observed. The enhanced CD8+T cell activity and Th1 immune responses by resveratrol administration might be related to the down-regulated PD-1 expression on pulmonary CD8+T cells and CD4+T cells. Resveratrol may also convert macrophages to M1 phenotype in the lungs of tumor bearing mice. However, it seems likely resveratrol has no effect on pulmonary myeloid-derived suppressor cell activation. Our results provide an evidence that resveratrol might be a promising candidate agent for adjuvant therapy in the process of TNBC metastasis.

IRF7 as an Immune-Related Prognostic Biomarker and Associated with Tumor Microenvironment in Low-Grade Glioma.

PurposeTumor microenvironment (TME) affects the occurrence and progression of low-grade glioma (LGG). The aim of this study is to identify TME-related genes that influence prognosis in LGG patients and to explore their function and role in tumor immunity.Patients and MethodsThe TME components of LGG samples in the Cancer Genome Atlas (TCGA) database were identified by the ESTIMATE method, and differentially expressed genes (DEGs) with significant differences in immune scores and stromal scores were screened out. The core genes of DEGs were screened out by protein–protein interaction (PPI) network. Furthermore, immune-related target genes significantly correlated with prognosis were identified. Survival analysis and correlation analysis showed the correlation between target genes and clinical features and prognosis. The expression differences of target genes were verified by external database Chinese Glioma Genome Atlas (CGGA). CIBERSORT software identified the proportion of tumor-infiltrating immune cells (TICs) that were significantly related to target genes. Gene set enrichment analysis (GSEA) could enrich the main functions related to high and low expression of target genes.ResultsA total of 1567 DEGs were screened out from 529 LGG samples in the TCGA database, and 146 immune-related genes affecting prognosis were found. A total of 403 core genes were obtained from PPI network. The target gene interferon regulatory factor 7 (IRF7) was significantly associated with prognosis and clinical features of the tumor. The CGGA database verified the relationship between high and low expression groups of IRF7 and prognosis. GSEA indicated that IRF7 was mainly enriched in immune-related activities, significantly correlated with T cells CD8, macrophages M1, macrophages M2 and monocytes.ConclusionThe IRF7 is involved in immune responses in TME of LGG, which in turn influenced tumor occurrence and progression. IRF7 can act as a potential biomarker for prognosis in patients with LGG and provide a target for tumor immunotherapy.

Combining IL-10 and Oncolytic Adenovirus Demonstrates Enhanced Antitumor Efficacy Through CD8+ T Cells.

Oncolytic viruses are of growing importance in cancer therapeutics since they combine direct oncolytic effect and the stimulation of antitumor immunity. Emerging evidences showed that the function of oncolytic viruses is dependent on immune response in tumor microenvironment, and the modulation of immunity could influence their efficacy. Here we combined the interleukin 10 (IL-10) and oncolytic adenovirus Ad-hTERT to treat lung cancer and explored the underlying mechanism under combination therapy. Lewis lung carcinoma (LLC) and B16F10 tumor-bearing immunocompetent C57BL/6 mice that received Ad-hTERT or IL-10 alone showed mild antitumor effect, while the combination therapy shrink tumor bulks and prolonged survival remarkably. In addition, IL-10 didn’t show direct influence on tumor cell viability or Ad-hTERT mediated tumor cell lysis in vitro. To further explore the influence of combination therapy mediated antitumor capacity, we eliminated CD8+ T, CD4+ T or natural killer (NK) cells in LLC and B16F10-bearing C57BL/6 mice, and found that CD8+ T cells were critical mediator in the combination therapy. The combination therapy induced intensive infiltration of CD8+ T cells in tumors, increased tumor-specific IFN-γ secretion by CD8+ T cells. The long-term tumor-specific immune memory induced by the combination therapy rejected rechallenge by respective tumor cell lines. This study demonstrated that the therapy combining IL-10 and Ad-hTERT augmented antitumor efficacy which was CD8+ T cells dependent. Our findings paved the way to combine cytokines and oncolytic viruses to enhance antitumor immunotherapy in treating cancer.

Development of an Oncogenic Driver Alteration Associated Immune-Related Prognostic Model for Stage I-II Lung Adenocarcinoma.

Lung adenocarcinoma (LUAD) needs to be stratified for its heterogeneity. Oncogenic driver alterations such as EGFR mutation, ALK translocation, ROS1 translocation, and BRAF mutation predict response to treatment for LUAD. Since oncogenic driver alterations may modulate immune response in tumor microenvironment that may influence prognosis in LUAD, the effects of EGFR, ALK, ROS1, and BRAF alterations on tumor microenvironment remain unclear. Immune-related prognostic model associated with oncogenic driver alterations is needed. In this study, we performed the Cox-proportional Hazards Analysis based on the L1-penalized (LASSO) Analysis to establish an immune-related prognostic model (IPM) in stage I-II LUAD patients, which was based on 3 immune-related genes (PDE4B, RIPK2, and IFITM1) significantly enriched in patients without EGFR, ALK, ROS1, and BRAF alterations in The Cancer Genome Atlas (TCGA) database. Then, patients were categorized into high-risk and low-risk groups individually according to the IPM defined risk score. The predicting ability of the IPM was validated in GSE31210 and GSE26939 downloaded from the Gene Expression Omnibus (GEO) database. High-risk was significantly associated with lower overall survival (OS) rates in 3 independent stage I-II LUAD cohorts (all P < 0.05). Moreover, the IPM defined risk independently predicted OS for patients in TCGA stage I-II LUAD cohort (P = 0.011). High-risk group had significantly higher proportions of macrophages M1 and activated mast cells but lower proportions of memory B cells, resting CD4 memory T cells and resting mast cells than low-risk group (all P < 0.05). In addition, the high-risk group had a significantly lower expression of CTLA-4, PDCD1, HAVCR2, and TIGIT than the low-risk group (all P < 0.05). In summary, we established a novel IPM that could provide new biomarkers for risk stratification of stage I-II LUAD patients.

Synergizing sunitinib and radiofrequency ablation to treat hepatocellular cancer by triggering the antitumor immune response

BackgroundMinimally invasive radiofrequency ablation (RFA) is used as a first-line treatment option for hepatocellular cancer (HCC) with the weaknesses of incomplete ablation, tumor recurrence, and inferior outcomes. To overcome this...

Decursin promotes HIF-1α proteasomal degradation and immune responses in hypoxic tumour microenvironment

BackgroundHypoxia and HIF-1α are important regulators of tumour growth and angiogenesis and could be attractive targets for cancer therapeutics. Decursin is an active compound extracted from the roots of Angelica gigas and has been shown to have potent anti-cancer and anti-angiogenic activities. However, whether decursin regulates HIF-1α activity and immune responses under hypoxic conditions is not yet understood. PurposeThe aim of this study was to identify whether decursin exhibits anti-cancer activity by targeting HIF-1α. Study designWe investigated whether decursin regulates HIF-1α protein stability and increases its degradation. In addition, we determined if decursin increases immune responses in tumour microenvironment to identify its hypoxia-associated anti-cancer activities. Materials and methodsWe performed the hypoxia-responsive element promoter–reporter assay, Western blot analysis, immune-fluorescence assay, semi-quantitative RT-PCR and ELISA for VEGF secretion, CCK-8 assay for cell proliferation, TUNEL assay for apoptosis and invasion assay in A549 human lung cancer or HCT116 human colon cancer cells. In vivo Lewis lung carcinoma (LLC) allograft mouse model was used to check tumour growth and immune responses in tumour microenvironment by immunohistochemistry analysis. ResultsWe observed that decursin inhibited HIF-1 activation under hypoxia by down-regulating the protein level of its subunit HIF-1α. It increased oxygen-dependant hydroxylation and ubiquitination of HIF-1α to promote HIF-1α degradation. Decursin also decreased mRNA expression of HIF-1α target genes. Decursin suppressed cancer cell proliferation, induced apoptosis and inhibited cancer cell invasion under hypoxia in cancer cells. In the allograft mouse tumour model, decursin reduced the hypoxic area and HIF-1α and PD-L1 expression. Infiltrating T cells (CD3+), helper T cells (CD4+) and cytotoxic (CD8+) T cells were accumulated, but regulatory T cells (Foxp3) and myeloid-derived suppressor cell-mediated immune suppressors (Arg1) were attenuated by decursin. ConclusionOur results suggest that decursin is a novel HIF-1α inhibitor that functions by promoting its proteasomal degradation and that it also helps improve T cell activation in tumour microenvironment; these findings provide new explanations about its anti-cancer and anti-angiogenic activity mechanisms.