Accumulation Of Immunosuppressive Cells Research Articles

Abstract IA005: Regional combating of immunosuppressive forces in the ovarian tumor microenvironment

Abstract IA005: Regional combating of immunosuppressive forces in the ovarian tumor microenvironment

Regulatory T cell-mediated immunosuppression orchestrated by cancer: towards an immuno-genomic paradigm for precision medicine.

Accumulating evidence indicates that aberrant signalling stemming from genetic abnormalities in cancer cells has a fundamental role in their evasion of antitumour immunity. Immune escape mechanisms include enhanced expression of immunosuppressive molecules, such as immune-checkpoint proteins, and the accumulation of immunosuppressive cells, including regulatory T (Treg) cells, in the tumour microenvironment. Therefore, Treg cells are key targets for cancer immunotherapy. Given that therapies targeting molecules predominantly expressed by Treg cells, such as CD25 or GITR, have thus far had limited antitumour efficacy, elucidating how certain characteristics of cancer, particularly genetic abnormalities, influence Treg cells is necessary to develop novel immunotherapeutic strategies. Hence, Treg cell-targeted strategies based on the particular characteristics of cancer in each patient, such as the combination of immune-checkpoint inhibitors with molecularly targeted agents that disrupt the immunosuppressive networks mediating Treg cell recruitment and/or activation, could become a new paradigm of cancer therapy. In this Review, we discuss new insights on the mechanisms by which cancers generate immunosuppressive networks that attenuate antitumour immunity and how these networks confer resistance to cancer immunotherapy, with a focus on Treg cells. These insights lead us to propose the concept of 'immuno-genomic precision medicine' based on specific characteristics of cancer, especially genetic profiles, that correlate with particular mechanisms of tumour immune escape and might, therefore, inform the optimal choice of immunotherapy for individual patients.

Orchestrated copper-based nanoreactor for remodeling tumor microenvironment to amplify cuproptosis-mediated anti-tumor immunity in colorectal cancer

Owing to the deficiency of infiltrating immune cells and the accumulation of immunosuppressive cells in tumor microenvironment (TME), immunotherapy remains enormous challenges in colorectal cancer (CRC). Here, CRC was found to be sensitive to cuproptosis which could further evoke immune responses and mediate immune resistance. Thus, a cuproptosis-mediated immunotherapy nanoreactor (CCJD-FA) was constructed by encapsulating copper-based shell-coated CaO2 and bromodomain-containing protein 4 inhibitor JQ-1 with DSPE-PEG-FA. Within tumor cells, CCJD-FA was disassembled under GSH environment, and the exposed CaO2 could generate H2O2 under the acidic condition, which reacted with the released Cu2+ via Fenton-like reaction to produce O2 for relieving hypoxia and Cu+ for inducing cuproptosis. Furthermore, CCJD-FA could also inhibit intracellular glycolysis and ATP generation, then blocking the Cu+ efflux protein ATP7B. Meanwhile, the IFN-γ-induced PD-L1 expression could be exactly reduced by the BRD4 suppression and oxygen supply. These effects sensitized cancer cells to cuproptosis, further evoking systemic immune responses and reshaping immunosuppressive TME to inhibit tumor growth.

Targeting the stimulator of interferon genes (STING) in breast cancer.

Breast cancer has a high occurrence rate globally and its treatment has demonstrated clinical efficacy with the use of systemic chemotherapy and immune checkpoint blockade. Insufficient cytotoxic T lymphocyte infiltration and the accumulation of immunosuppressive cells within tumours are the primary factors responsible for the inadequate clinical effectiveness of breast cancer treatment. The stimulator of interferon genes (STING) represents a pivotal protein in the innate immune response. Upon activation, STING triggers the activation and enhancement of innate and adaptive immune functions, resulting in therapeutic benefits for malignant tumours. The STING signalling pathway in breast cancer is influenced by various factors such as deoxyribonucleic acid damage response, tumour immune microenvironment, and mitochondrial function. The use of STING agonists is gaining momentum in breast cancer research. This review provides a comprehensive overview of the cyclic guanosine monophosphate-adenosine monophosphate synthase-STING pathway, its agonists, and the latest findings related to their application in breast cancer.

IL-6 Levels Correlate with Prognosis and Immunosuppressive Stromal Cells in Patients with Colorectal Cancer.

The prognosis for patients with colorectal cancer (CRC) is determined by tumor characteristics as well as the host immune response. This study investigated the relationship between an immunosuppressive state and patient prognosis by evaluating the systemic and tumor microenvironment (TME) interleukin (IL)-6 levels. Preoperative serum IL-6 levels were measured using an electrochemiluminescence assay. Expression of IL-6 in tumor and stromal cells was evaluated immunohistochemically in 209 patients with resected CRC. Single-cell analysis of tumor-infiltrating immune cells was performed using mass cytometry in 10 additional cases. Elevated serum IL-6 levels were associated with elevated stromal IL-6 levels and a poor prognosis for patients with CRC. High IL-6 expression in stromal cells was associated with low-density subsets of CD3+ and CD4+ T cells as well as FOXP3+ cells. Mass cytometry analysis showed that IL-6+ cells among tumor-infiltrating immune cells were composed primarily of myeloid cells and rarely of lymphoid cells. In the high-IL-6-expression group, the percentages of myeloid-derived suppressor cells (MDSCs) and CD4+FOXP3highCD45RA- effector regulatory T cells (eTreg) were significantly higher than in the low-IL-6-expression group. Furthermore, the proportion of IL-10+ cells in MDSCs and that of IL-10+ or CTLA-4+ cells in eTregs correlated with IL-6 levels. Elevated serum IL-6 levels were associated with stromal IL-6 levels in CRC. High IL-6 expression in tumor-infiltrating immune cells also was associated with accumulation of immunosuppressive cells in the TME.

Fecal microbiota transplantation inhibits colorectal cancer progression: Reversing intestinal microbial dysbiosis to enhance anti-cancer immune responses.

Many lines of evidence demonstrate the associations of colorectal cancer (CRC) with intestinal microbial dysbiosis. Recent reports have suggested that maintaining the homeostasis of microbiota and host might be beneficial to CRC patients, but the underlying mechanisms remain unclear. In this study, we established a CRC mouse model of microbial dysbiosis and evaluated the effects of fecal microbiota transplantation (FMT) on CRC progression. Azomethane and dextran sodium sulfate were used to induce CRC and microbial dysbiosis in mice. Intestinal microbes from healthy mice were transferred to CRC mice by enema. The vastly disordered gut microbiota of CRC mice was largely reversed by FMT. Intestinal microbiota from normal mice effectively suppressed cancer progression as assessed by measuring the diameter and number of cancerous foci and significantly prolonged survival of the CRC mice. In the intestine of mice that had received FMT, there were massive infiltration of immune cells, including CD8+ T and CD49b+ NK, which is able to directly kill cancer cells. Moreover, the accumulation of immunosuppressive cells, Foxp3+ Treg cells, seen in the CRC mice was much reduced after FMT. Additionally, FMT regulated the expressions of inflammatory cytokines in CRC mice, including down-regulation of IL1a, IL6, IL12a, IL12b, IL17a, and elevation of IL10. These cytokines were positively correlated with Azospirillum_sp._47_25, Clostridium_sensu_stricto_1, the E. coli complex, Akkermansia, Turicibacter, and negatively correlated with Muribaculum, Anaeroplasma, Candidatus_Arthromitus, and Candidatus Saccharimonas. Furthermore, the repressed expressions of TGFb, STAT3 and elevated expressions of TNFa, IFNg, CXCR4 together promoted the anti-cancer efficacy. Their expressions were positively correlated with Odoribacter, Lachnospiraceae-UCG-006, Desulfovibrio, and negatively correlated with Alloprevotella, Ruminococcaceae UCG-014, Ruminiclostridium, Prevotellaceae UCG-001 and Oscillibacter. Our studies indicate that FMT inhibits the development of CRC by reversing gut microbial disorder, ameliorating excessive intestinal inflammation and cooperating with anti-cancer immune responses.

Single-cell transcriptomics reveals shared immunosuppressive landscapes of mouse and human neuroblastoma

BackgroundHigh-risk neuroblastoma is a pediatric cancer with still a dismal prognosis, despite multimodal and intensive therapies. Tumor microenvironment represents a key component of the tumor ecosystem the complexity of which...

Inhibition of TFEB promotes tumor-educated dendritic cells activation to enhance antitumor immune responses

Tumors can induce the generation and accumulation of immunosuppressive cells in –the tumor microenvironment (TME). Among them, tumor-educated dendritic cells (TEDCs) involved in tolerance induction contribute greatly to the progression of tumors. However, the mechanisms governing the immunosuppressive function of dendritic cells in the TME are unclear. In this study, we found that the expression of transcription factor EB (TFEB) was significantly increased in TEDCs induced by cancer cell supernatant. TFEB knockdown significantly promoted the differentiation and maturation of TEDCs, with upregulated expression of CD11c and costimulatory molecules (CD86 and MHC-II) but reduced expression of the inhibitory molecule PD-L1, and enhanced their ability to induce Th1 proliferation and differentiation. Moreover, TEDCs with TFEB knockdown significantly reduced tumor growth with increased infiltration of CD11c+MHC-II+ dendritic cells and effector T cells in tumor masses, thus leading to a delay in tumor progression. These findings demonstrate a critical role of TFEB in regulating the immunosuppression of TEDCs, with potential implications as an antitumor immune therapeutic approach.

Lenvatinib- and vadimezan-loaded synthetic high-density lipoprotein for combinational immunochemotherapy of metastatic triple-negative breast cancer

Metastatic triple-negative breast cancer (TNBC) is the most aggressive type of breast cancer. Combination of systemic chemotherapy and immune checkpoint blockade is effective but of limited benefit due to insufficient intratumoral infiltration of cytotoxic T lymphocytes (CTLs) and the accumulation of immunosuppressive cells. Herein, we designed a lenvatinib- and vadimezan-loaded synthetic high-density lipoprotein (LV-sHDL) for combinational immunochemotherapy of metastatic TNBC. The LV-sHDL targeted scavenger receptor class B type 1-overexpressing 4T1 cells in the tumor after intravenous injection. The multitargeted tyrosine kinase inhibitor (TKI) lenvatinib induced immunogenic cell death of the cancer cells, and the stimulator of interferon genes (STING) agonist vadimezan triggered local inflammation to facilitate dendritic cell maturation and antitumor macrophage differentiation, which synergistically improved the intratumoral infiltration of total and active CTLs by 33- and 13-fold, respectively. LV-sHDL inhibited the growth of orthotopic 4T1 tumors, reduced pulmonary metastasis, and prolonged the survival of animals. The efficacy could be further improved when LV-sHDL was used in combination with antibody against programmed cell death ligand 1. This study highlights the combination use of multitargeted TKI and STING agonist a promising treatment for metastatic TNBC.

Breast Cancer Tumor Microenvironment and Molecular Aberrations Hijack Tumoricidal Immunity.

Simple SummaryImmune therapy is designed to stimulate tumoricidal effects in a variety of solid tumors including breast carcinomas. However, the emergence of resistant clones leads to treatment failure. Understanding the molecular, cellular, and microenvironmental aberrations is crucial to uncovering underlying mechanisms and developing advanced strategies for preventing or combating these resistant malignancies. This review will summarize research findings revealing various mechanisms employed to hijack innate and adaptive immune surveillance mechanisms, develop hypoxic and tumor promoting metabolism, and foster an immune tolerance microenvironment. In addition, it will highlight potential targets for therapeutic approaches.Breast cancer is the most common malignancy among females in western countries, where women have an overall lifetime risk of >10% for developing invasive breast carcinomas. It is not a single disease but is composed of distinct subtypes associated with different clinical outcomes and is highly heterogeneous in both the molecular and clinical aspects. Although tumor initiation is largely driven by acquired genetic alterations, recent data suggest microenvironment-mediated immune evasion may play an important role in neoplastic progression. Beyond surgical resection, radiation, and chemotherapy, additional therapeutic options include hormonal deactivation, targeted-signaling pathway treatment, DNA repair inhibition, and aberrant epigenetic reversion. Yet, the fatality rate of metastatic breast cancer remains unacceptably high, largely due to treatment resistance and metastases to brain, lung, or bone marrow where tumor bed penetration of therapeutic agents is limited. Recent studies indicate the development of immune-oncological therapy could potentially eradicate this devastating malignancy. Evidence suggests tumors express immunogenic neoantigens but the immunity towards these antigens is frequently muted. Established tumors exhibit immunological tolerance. This tolerance reflects a process of immune suppression elicited by the tumor, and it represents a critical obstacle towards successful antitumor immunotherapy. In general, immune evasive mechanisms adapted by breast cancer encompasses down-regulation of antigen presentations or recognition, lack of immune effector cells, obstruction of anti-tumor immune cell maturation, accumulation of immunosuppressive cells, production of inhibitory cytokines, chemokines or ligands/receptors, and up-regulation of immune checkpoint modulators. Together with altered metabolism and hypoxic conditions, they constitute a permissive tumor microenvironment. This article intends to discern representative incidents and to provide potential innovative therapeutic regimens to reinstate tumoricidal immunity.

Abstract 2720: Perivascular accumulation of immunosuppressive cells in the stroma of human triple negative breast carcinomas: implications for immunotherapy

Abstract A distinct subset of tumor-associated macrophages (TAMs) accumulate around blood vessels in mouse tumors where they stimulate tumor angiogenesis and various steps in the metastatic pathway. They also limit tumor responses to frontline anti-cancer therapies like irradiation and chemotherapy, and dampen anti-tumor immunity by recruiting regulatory T cells and suppressing the proliferation of T cells. However, the presence and phenotype of such perivascular (PV) TAMs have yet to be investigated in human tumors. In this study we have used MultiOmyx multiplex immunofluorescence coupled to advanced analytics to compare the distribution and phenotype of TAMs, CD4+ and CD8+ T cells, and CD4+FOXP3+ regulatory T cells (Tregs) in PV vs non-PV areas in the stroma and tumor cell islands (TCIs) of 40 human triple negative breast carcinomas (TNBCs) - and whether this was altered after neoadjuvant chemotherapy. Tregs along with distinct subsets of TAMs and CD4+ and CD8+ T cells were found to preferentially accumulate in PV areas (ie. within 50um of CD31+ blood vessels), especially in the tumor stroma. CD163+ TIM3+ TAMs often made direct contact with the abluminal surface of blood vessels in these sites, and at increased numbers in in chemotherapy-treated TNBCs. This suggests they may regulate the formation/function of blood vessels in relapsing tumors. In both untreated and chemotherapy-treated tumors, a major subset of CD163+ TAMs lacked PDL1 expression and also accumulated preferentially in stromal PV areas, where many made direct contact with PD1- CD4+ or PD1- CD8+ T cells as well as FOXP3+CD4+ Tregs. We are currently investigating whether such close contact of these subsets of T cells with two immunosuppressive cell types (ie. CD163+ TAMs and Tregs) inhibits their subsequent anti-tumor functions. Taken together, our study shows that, as T cells cross blood vessels into TNBCs, they encounter a high density of at least two immunosuppressive cell types in the perivascular niche. This could lead to the inactivation of T cells before they migrate further into tumors, thereby reducing anti-tumor immunity. It could also limit the efficacy of cancer immunotherapies mediated by activated T cells. Identifying and targeting the mechanism causing these suppressive cells to accumulate and interact in such sites could remove this inhibition. Citation Format: Mohammed Ridha Moamin, Richard J. Allen, Nicholas Stavrou, Mate Nagy, Anna Juncker-Jensen, Claire E. Lewis. Perivascular accumulation of immunosuppressive cells in the stroma of human triple negative breast carcinomas: implications for immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2720.

Expression of a constitutively active p38α mutant in mice causes early death, anemia, and accumulation of immunosuppressive cells.

The MAP kinase p38α is associated with numerous processes in eukaryotes, and its elevated activity is a prominent feature of inflammatory diseases, allergies, and aging. Since p38α is a nodal component of a complex signaling network, it is difficult to reveal exactly how p38α contributes to disparate outcomes. Identification of p38α -specific effects requires activation of p38α perse invivo. We generated a transgenic mouse model that meets this requirement by allowing inducible and reversible expression of an intrinsically active p38α molecule (p38αD176A+F327S ). p38α's activation across all murine tissues resulted in a significant loss of body weight and death of about 40% of the mice within 17weeks of activation, although most tissues were unaffected. Flow cytometric analysis of the lungs and bronchoalveolar lavage fluid detected an accumulation of 'debris' within the airways, suggesting impaired clearance. It also revealed increased numbers of alternatively activated alveolar macrophages and myeloid-derived suppressor cells within the lung, pointing at suppression and resolution of inflammation. Blood count suggested that mice expressing p38αD176A+F327S suffer from hemolytic anemia. Flow cytometry of bone marrow revealed a reduced number of hematopoietic stem cells and abnormalities in the erythroid lineage. Unexpectedly, p38α's substrate MAPKAPK2, mitogen-activated protein kinase-activated protein kinase 2 was downregulated in mice expressing p38αD176A+F327S , suggesting that constitutive activity of p38α may impose pathological phenotypes by downregulating downstream components, perhaps via a feedback inhibition mechanism. In summary, this new mouse model shows that induced p38α activity perse is hazardous to mouse vitality and welfare, although pathological parameters are apparent only in blood count, bone marrow, and lungs.

Exogenous interleukin-33 promotes hepatocellular carcinoma growth by remodelling the tumour microenvironment

BackgroundInterleukin-33 (IL-33) is an effective inducer of pro-inflammatory cytokines regulating innate and adaptive immunity. Inflammation could be a double-edged sword, promoting or inhibiting tumour growth. To date, the roles and mechanisms of IL-33 in tumours remain controversial. Here, we examined the effect of exogenous IL-33 on the biological characteristics of hepatocellular carcinoma (HCC) and the possible mechanism of action.MethodsIn this study, IL-33 expression in the tissues of 69 HCC patients was detected and its relationship with prognosis was evaluated. After establishing a mouse HCC model and IL-33 treatment operation, the infiltration of splenic myeloid-derived suppressor (MDSCs), dendritic (DCs), regulatory T, and natural killer (NK) cells was detected by flow cytometry analysis, and the vascular density of the tumour tissues was detected by immunohistochemistry to reveal the mechanism of IL-33 in HCC proliferation. Finally, the Cancer Genome Atlas database was used to analyse Gene Ontology terms the and Kyoto Encyclopaedia of Genes and Genomes pathway. Moreover, the chi-square test, two-tailed unpaired Student’s t-test, and multiple t-tests were performed using SPSS version 23.0 and GraphPad Prism 8.0 software.ResultsThe IL-33 expression level was negatively correlated with the overall survival of HCC patients, suggesting its potential clinical significance in the prognosis of HCC. We found that systemic IL-33 administration significantly promoted the tumour size in vivo. Furthermore, the IL-33-treated mice presented decreased frequencies of tumouricidal NK and CD69+ CD8+ T cells. After IL-33 treatment, the incidence of monocytic MDSCs and conventional DCs increased, while that of granulocytic MDSCs decreased. Moreover, IL-33 promoted the formation of intracellular neovascularization. Therefore, IL-33 accelerated HCC progression by increasing the accumulation of immunosuppressive cells and neovascularization formation. Finally, we found that the transcription of IL-33 was closely related to the PI3K-Akt and MAPK pathways in Gene Set Enrichment Analysis plots, which were involved in the tumourigenesis and pathogenesis of HCC.ConclusionsTaken together, IL-33 may be a key tumour promoter of HCC proliferation and tumourigenicity, an important mediator, and a potential therapeutic target for regulating HCC progression.

Tumor RIPK1 signaling mediates intrinsic and extrinsic mechanisms of resistance to immune checkpoint blockade therapy

Abstract Immune checkpoint blockade (ICB) therapy can result in impressive clinical responses of multiple cancer types, however resistance is common. Mechanisms of resistance include both tumor intrinsic evasion of immune mediated killing and tumor extrinsic features in the immune-microenvironment, such as poor infiltration of tumor reactive T cells and accumulation of immunosuppressive cells. By examining transcriptomic features associated with response or resistance to ICB, we found the expression of Receptor Interacting Protein Kinase 1 (RIPK1), a critical regulator of inflammation and cell death, is significantly higher in ICB resistant murine tumor cells. We hypothesized that RIPK1 expression in the tumor cells may influence both intrinsic and extrinsic mechanisms regulating response to ICB. To test this hypothesis, we genetically deleted RIPK1 in murine tumor cell lines and found improved overall survival upon treatment compared to tumors expressing RIPK1. Mechanistically, deletion of RIPK1 in the tumor cells results in favorable changes in the tumor microenvironment, including an increased frequency of effector T cells, a decreased frequency of immunosuppressive myeloid cells, and changes in inflammatory cytokine secretion that can impact these changes in immune infiltrate. Further, deletion of the apoptosis effector Caspase 8 reversed RIPK1 KO tumor sensitization to ICB, suggesting that tumor sensitization to cell death also contributes to improved response to ICB upon RIPK1 deletion. In conclusion, RIPK1 is an important regulator at the intersection of tumor intrinsic and extrinsic pathways governing response to ICB, and therefore is a potential target for combinatorial therapies to improve patient response to ICB.

Deciphering and Reversing Immunosuppressive Cells in the Treatment of Hepatocellular Carcinoma.

Use of immune checkpoint inhibitors (ICIs) in hepatocellular carcinoma (HCC) has been partially successful. However, most HCC patients do not respond to immunotherapy. HCC has been shown to induce several immune suppressor mechanisms in patients. These suppressor mechanisms include involvement of myeloid-derived suppressor cells, regulatory T-cells, functionally impaired dendritic cells (DCs), neutrophils, monocytes, and tumor associated macrophages. The accumulation of immunosuppressive cells may lead to an immunosuppressive tumor microenvironment as well as the dense fibrotic stroma which may contribute to immune tolerance. Our laboratory has been investigating different cellular mechanisms of immune suppression in HCC patients. In vitro as well as in vivo studies have demonstrated that abrogation of the suppressor cells enhances or unmasks tumor-specific antitumor immune responses. Two or three effective systemic therapies including ICIs and/or molecular targeted therapies and the addition of innovative combination therapies targeting immune suppressor cells may lead to increased immune recognition with a greater tumor response. We reviewed the literature for the latest research on immune suppressor cells in HCC, and here we provide a comprehensive summary of the recent studies in this field.

Phenotype molding of T cells in colorectal cancer by single-cell analysis.

The majority of patients with microsatellite stable (MSS) colorectal cancer (CRC) do not benefit from the immunotherapies directed at rescuing T-cell functions. Therefore, complete understanding of T-cell phenotypes and functional status in the CRC microenvironment is desirable. Here, we applied single-cell mass cytometry to mold the T-cell phenotype in 18 patients with MSS CRC for better understanding of CRC as a systemic disease and to search for tumor-driven T-cell profile changes. We show interpatient and intrapatient phenotypic diversity of T-cell subsets. We revealed increased immunosuppressive/exhausted T-cell phenotypes at tumor lesions. CD8+ CD28- immunosenescent T cells with impaired proliferation capacity dominate the T-cell compartment. As per the transcriptome and quantitative real time-PCR analysis, the accumulation of immunosuppressive cells is driven by the tumor microenvironment. T-cell profiles are similar between patients at early and late stages, indicating that the immunosuppressive microenvironment is formulated early during CRC development. Mapping of T-cell infiltration and understanding of the mechanisms underlying their regulation may provide valuable information to boost the immune response in patients with MSS CRC.

Contributions of T cell dysfunction to the resistance against anti-PD-1 therapy in oral carcinogenesis

BackgroundProgrammed death 1 (PD-1) blockade has great effect in the prevention of oral precancerous lesions, but the drug resistance has also been observed. The determinants of immune resistance during the malignant transformation are poorly understood.MethodsAnti-PD-1 antibody was administered in the 4NQO-induced carcinogenesis mouse models. The mice were then subdivided into PD-1 resistance(PD-1R) group and PD-1 sensitive(PD-1S) group according to the efficacy. The expression of PD-1 and PD-L1, and the abundance of CD3+ T cells in tumor microenvironment between the two groups was tested by immunohistochemistry. In addition, the activation and effector functions, as well as the accumulation of immunosuppressive cells and expression of immune checkpoints of T cells in the draining lymph nodes and spleen between PD-1R and PD-1S group were analyzed by flow cytometry.ResultsOur results showed that T cell infiltration in tumor microenvironment, effector T cell cytokine secretion and central memory T cell accumulation in peripheral lymphoid organs were all inhibited in the anti-PD-1 resistance group. Furthermore, we found that an increase of regulatory T cell (Treg) population contributed to the resistance of the anti-PD-1 therapy. Notably, TIM-3 was found to be the only immunosuppressive molecule that mediated the resistance to anti-PD-1 therapy in the oral malignant transformation model.ConclusionsOur findings identified a novel mechanism that T cell dysfunction contributes to the immune resistance during the malignant transformation of the oral mucosa. This study provides new targets for improving the efficacy of immunotherapy for early stage of tumorigenesis.

Chemotherapy alters the increased numbers of myeloid-derived suppressor and regulatory T cells in children with acute lymphoblastic leukemia

Introduction: Acute lymphoblastic leukemia (ALL) is the most common cancer diagnosed in children. The precise mechanism behind the relapse in this disease is not clearly known. One possible mechanism could be the accumulation of immunosuppressive cells, including myeloid-derived suppressor cells (MDSCs) and T regulatory cells (Tregs) which we and others have reported to mediate suppression of anti-tumor immune responses.Aim: In this study, we aimed to analyze the numbers of these cells in a population of B-ALL pediatric patients.Methods: Peripheral blood samples withdrawn from B-ALL pediatric patients (n = 45 before, during and after the induction phase of chemotherapy. Using multi parametric flow cytometric analysis. MDSCs were identified as Lin–HLA-DR–CD33+CD11b+; and Treg cells were defined as CD4+CD25+CD127–/low.Results: Early diagnosed B-ALL patients showed significant increases in the numbers of MDSCs and Tregs as compared to healthy volunteers. During induction of chemotherapy, however, the patients showed higher and lower numbers of MDSCs and Treg cells, respectively as compared to early diagnosed patients (i.e., before chemotherapy). After induction of chemotherapy, the numbers of MDSCs and Treg cells showed higher increases and decreases, respectively as compared to the numbers in patients during chemotherapy.Conclusion: Our results indicate that B-ALL patients harbor high numbers of both MDSCs and Tregs cells. This pilot study opens a new avenue to investigate the mechanism mediating the emergence of these cells on larger number of B-ALL patients at different treatment stages.

Correlation of VEGF with immune suppression involving MDSC, malnutrition, and prognosis in patients with gastric and colorectal cancer.

582 Background: Although a causal relationship for inflammation and immunity of cancer is more widely accepted today, the precise cell mechanisms mediating this relationship have not been elucidated. Vascular endothelial growth factor (VEGF), previously known as vascular permeability factor. 45 kDa protein, belongs to a family of platelet-derived growth factors. VEGF, inflammation-related protein, could contribute to the accumulation of immunosuppressive cells (MDSC, Treg, TAM, and Tie-2-expressingmonocytes) in tumor-bearing hosts through direct or indirect mechanisms. Methods: We tested the serum levels of VEGF by ELISA in 106 patients including 43 with gastric and 63 with colorectal cancer, and they were increased in advanced stages of gastric and colorectal cancer. Production of IL-17, pro-inflammatory cytokine, with a stimulation of PHA was measured by ELISA and MDSC (myeloid-derived suppressor cells), one of major immunosuppressing cells, was measured by flow cytometry (CD11b+CD14-CD33+). Neutrophil to lymphocyte ratio (NLR) and C-reactive protein (CRP) were used as inflammatory markers. Production of IL-12 and SI (stimulation index) of blastogenic response of lymphocytes were used as markers of cell-mediated immune response. Results: The concentrations of VEGF were positively correlated with levels of MDSC, production of IL-17, NLR and CRP, and were inversely correlated with IL-12 production, SI and nutritional markers including prealbumin and retinol binding protein. The patients were both divided into two groups with a serum level of VEGF (330 pg/ml) and OS (overall survival) of patients with stages III and IV gastric or colorectal cancer were both significantly worse in patients with high levels of VEGF than in those with low VEGF although the differences were not significant in patients with stagesⅠand II. Conclusions: The results of the present study suggested that VEGF may have an impact on advancement and progression involving inflammation and serve as useful markers of the immune suppression involving MDSC, malnutrition and poor prognosis in patients with gastric and colorectal cancer.

Abstract 1017: Lipid metabolic reprogramming drives resistance to PD1 blockage

Abstract The mechanisms underlying immunosuppression and resistance to PD1 inhibitors in cancer are not well understood. We attempted to fill this gap with an integrated analysis of mRNA, microRNA, and protein expression in an anti-PD1-resistant lung adenocarcinoma mouse model. The model was created by in vivo passage of 344SQ murine lung cancer cells (p53R172HΔg/+K-rasLA1/+) in a syngeneic host repeatedly dosed with anti-mouse PD1 antibodies. Anti-PD1-resistant 344SQ (344SQ_R) and 344SQ parental (344SQ_P) cells were then inoculated into syngeneic 129Sv/ev mice, which were then dosed twice with anti-PD1 or control IgG antibodies. Tumor tissues were collected and analyzed as follows: transcriptome with Affymetrix; protein levels by reverse phase protein array analysis; signature enrichment by gene set enrichment analysis; metabolome by mass spectrometry; and lipid content with fluorescent probes Oil O rad and BODIPY. We also isolated tumor-infiltrating immune cells for flow cytometry and gene expression analyses. We identified lipid-related metabolic pathways as being the most highly enriched in anti-PD1-resistant tumors (344SQ_R) vs. their 344SQ_P counterparts; the resistant cells also had more lipid droplets than the 344SQ_P cells. The anti-PD1-resistant tumors overexpressed several genes involved in lipogenesis and fatty acid pathways (e.g., fatty acid binding proteins [FABPs], fatty acid synthase, acetyl-coA-acyltransferase 2, fatty acid elongases). Specifically, FABP overexpression promoted fatty acid uptake and lipid-droplet accumulation in resistant tumors. Lipid-sensitive targets linked to inflammation and insulin signaling (e.g,. stress-activated kinases such as JNK and NFκB) were altered in 344SQ_R vs. 344SQ_P tumors. Mechanistically, JNK downregulation by NFκB-regulated microRNAs protected PD1-resistant tumors from lipotoxicity caused by FABPs upregulation and fatty acid uptake. FABP levels were higher in plasma from 344SQ_R than from 344SQ_P tumors. Tumor-infiltrating macrophages from 344SQ_R tumors had 4 times the amount of FABP mRNA than parental tumors and a correspondingly higher percentage of M2-like macrophages. 344SQ_R tumors promoted immune suppressive cells by upregulating FABPs expression in M2-like macrophages, marked by increased fatty acid intake and fatty acid oxidation. Conversely, percentages of CD4+ and CD8+ tumor-infiltrating lymphocytes were reduced in the resistant tumors. These results suggest that lipid metabolic rewiring drives resistance PD1 inhibitors supporting the accumulation of immunosuppressive cells, including M2-like macrophages, preventing type I immune responses elicited by T cells. Collectively, these findings reveal new potential lipid-related targets for drug development or new treatments combining inhibitors of these targets with anti-PD1 therapy. Citation Format: Maria A. Cortez, Sharareh Niknam, Efrosini Cuko, Jonathan E. Schoenhals, Hampartsoum Barsoumian, Ahmed I. Younes, Ailin Li, Jody V. Vykoukal, Cristina Ivan, George A. Calin, Patrick Hwu, James W. Welsh. Lipid metabolic reprogramming drives resistance to PD1 blockage [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1017. doi:10.1158/1538-7445.AM2017-1017