ABSTRACT
Type I interferon (IFN) is a class of proinflammatory cytokines with a dual role on malignant transformation, tumor progression, and response to therapy. On the one hand, robust, acute, and resolving type I IFN responses have been shown to mediate prominent anticancer effects, reflecting not only their direct cytostatic/cytotoxic activity on (at least some) malignant cells, but also their pronounced immunostimulatory functions. In line with this notion, type I IFN signaling has been implicated in the antineoplastic effects of various immunogenic therapeutics, including (but not limited to) immunogenic cell death (ICD)-inducing agents and immune checkpoint inhibitors (ICIs). On the other hand, weak, indolent, and non-resolving type I IFN responses have been demonstrated to support tumor progression and resistance to therapy, reflecting the ability of suboptimal type I IFN signaling to mediate cytoprotective activity, promote stemness, favor tolerance to chromosomal instability, and facilitate the establishment of an immunologically exhausted tumor microenvironment. Here, we review fundamental aspects of type I IFN signaling and their context-dependent impact on malignant transformation, tumor progression, and response to therapy.
Subject(s)
Antineoplastic Agents , Interferon Type I , Neoplasms , Humans , Neoplasms/drug therapy , Antineoplastic Agents/pharmacology , Antineoplastic Agents/therapeutic use , Cytokines , Tumor MicroenvironmentABSTRACT
In some settings, cancer cells responding to treatment undergo an immunogenic form of cell death that is associated with the abundant emission of danger signals in the form of damage-associated molecular patterns. Accumulating preclinical and clinical evidence indicates that danger signals play a crucial role in the (re-)activation of antitumor immune responses in vivo, thus having a major impact on patient prognosis. We have previously demonstrated that the presence of calreticulin on the surface of malignant blasts is a positive prognostic biomarker for patients with acute myeloid leukemia (AML). Calreticulin exposure not only correlated with enhanced T-cell-dependent antitumor immunity in this setting but also affected the number of circulating natural killer (NK) cells upon restoration of normal hematopoiesis. Here, we report that calreticulin exposure on malignant blasts is associated with enhanced NK cell cytotoxic and secretory functions, both in AML patients and in vivo in mice. The ability of calreticulin to stimulate NK-cells relies on CD11c+CD14high cells that, upon exposure to CRT, express higher levels of IL-15Rα, maturation markers (CD86 and HLA-DR) and CCR7. CRT exposure on malignant blasts also correlates with the upregulation of genes coding for type I interferon. This suggests that CD11c+CD14high cells have increased capacity to migrate to secondary lymphoid organs, where can efficiently deliver stimulatory signals (IL-15Rα/IL-15) to NK cells. These findings delineate a multipronged, clinically relevant mechanism whereby surface-exposed calreticulin favors NK-cell activation in AML patients.
Subject(s)
Calreticulin , Leukemia, Myeloid, Acute , Animals , Calreticulin/genetics , Calreticulin/metabolism , Cytotoxicity, Immunologic , Humans , Interleukin-15 , Killer Cells, Natural , Leukemia, Myeloid, Acute/therapy , Lymphocyte Activation , MiceABSTRACT
Adoptive cell therapy (ACT) is becoming a prominent alternative therapeutic treatment for cancer patients relapsing on traditional therapies. In parallel, antibodies targeting immune checkpoint molecules, such as cytotoxic-T-lymphocyte-associated antigen 4 (CTLA-4) and cell death protein 1 pathway (PD-1), are rapidly being approved for multiple cancer types, including as first line therapy for PD-L1-expressing non-small-cell lung cancer. The combination of ACT and checkpoint blockade could substantially boost the efficacy of ACT. In this study, we generated a novel self-delivering small interfering RNA (siRNA) (sdRNA) that knocked down PD-1 expression on healthy donor T cells as well as patient-derived tumor-infiltrating lymphocytes (TIL). We have developed an alternative chemical modification of RNA backbone for improved stability and increased efficacy. Our results show that T cells treated with sdRNA specific for PD-1 had increased interferon γ (IFN-γ) secreting capacity and that this modality of gene expression interference could be utilized in our rapid expansion protocol for production of TIL for therapy. TIL expanded in the presence of PD-1-specific sdRNA performed with increased functionality against autologous tumor as compared to control TIL. This method of introducing RNAi into T cells to modify the expression of proteins could easily be adopted into any ACT protocol and will lead to the exploration of new combination therapies.
Subject(s)
Lung Neoplasms/metabolism , Lung Neoplasms/therapy , Melanoma/immunology , Melanoma/therapy , Programmed Cell Death 1 Receptor/metabolism , T-Lymphocytes/metabolism , Cell- and Tissue-Based Therapy/methods , Flow Cytometry , HeLa Cells , Humans , Immunotherapy, Adoptive/methods , Interferon-gamma/genetics , Interferon-gamma/metabolism , Lung Neoplasms/immunology , Melanoma/metabolism , Programmed Cell Death 1 Receptor/genetics , RNA Interference/physiologyABSTRACT
Cancer cell death can be perceived as immunogenic by the host only when malignant cells emit immunostimulatory signals (so-called "damage-associated molecular patterns," DAMPs), as they die in the context of failing adaptive responses to stress. Accumulating preclinical and clinical evidence indicates that the capacity of immunogenic cell death to (re-)activate an anticancer immune response is key to the success of various chemo- and radiotherapeutic regimens. Malignant blasts from patients with acute myeloid leukemia (AML) exposed multiple DAMPs, including calreticulin (CRT), heat-shock protein 70 (HSP70), and HSP90 on their plasma membrane irrespective of treatment. In these patients, high levels of surface-exposed CRT correlated with an increased proportion of natural killer cells and effector memory CD4+ and CD8+ T cells in the periphery. Moreover, CRT exposure on the plasma membrane of malignant blasts positively correlated with the frequency of circulating T cells specific for leukemia-associated antigens, indicating that ecto-CRT favors the initiation of anticancer immunity in patients with AML. Finally, although the levels of ecto-HSP70, ecto-HSP90, and ecto-CRT were all associated with improved relapse-free survival, only CRT exposure significantly correlated with superior overall survival. Thus, CRT exposure represents a novel powerful prognostic biomarker for patients with AML, reflecting the activation of a clinically relevant AML-specific immune response.
Subject(s)
Blast Crisis/immunology , Blast Crisis/pathology , Calreticulin/metabolism , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/immunology , Alarmins/metabolism , CD8-Positive T-Lymphocytes/immunology , Cell Death , Female , Gene Expression Profiling , Gene Expression Regulation, Leukemic , HSP70 Heat-Shock Proteins/metabolism , HSP90 Heat-Shock Proteins/metabolism , Humans , Immunity , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/metabolism , Male , Middle Aged , Multivariate Analysis , Phenotype , Proportional Hazards Models , Th1 Cells/immunology , Transcription, Genetic , Treatment OutcomeABSTRACT
Dendritic cell (DC) vaccines have been demonstrated to elicit immunological responses in numerous cancer immunotherapy trials. However, long-lasting clinical effects are infrequent. We therefore sought to establish a protocol to generate DC with greater immunostimulatory capacity. Immature DC were generated from healthy donor monocytes by culturing in the presence of IL-4 and GM-CSF and were further differentiated into mature DC by the addition of cocktails containing different cytokines and toll-like receptor (TLR) agonists. Overall, addition of IFNγ and the TLR7/8 agonist R848 during maturation was essential for the production of high levels of IL-12p70 which was further augmented by adding the TLR3 agonist poly I:C. In addition, the DC matured with IFNγ, R848, and poly I:C also induced upregulation of several other pro-inflammatory and Th1-skewing cytokines/chemokines, co-stimulatory receptors, and the chemokine receptor CCR7. For most cytokines and chemokines the production was even further potentiated by addition of the TLR4 agonist LPS. Concurrently, upregulation of the anti-inflammatory cytokine IL-10 was modest. Most importantly, DC matured with IFNγ, R848, and poly I:C had the ability to activate IFNγ production in allogeneic T cells and this was further enhanced by adding LPS to the cocktail. Furthermore, epitope-specific stimulation of TCR-transduced T cells by peptide- or whole tumor lysate-loaded DC was efficiently stimulated only by DC matured in the full maturation cocktail containing IFNγ and the three TLR ligands R848, poly I:C, and LPS. We suggest that this cocktail is used for future clinical trials of anti-cancer DC vaccines.
Subject(s)
Dendritic Cells/immunology , Interferon-gamma/pharmacology , T-Lymphocytes/immunology , Toll-Like Receptors/agonists , Cell Differentiation , HumansABSTRACT
Recent studies have identified molecular events characteristic of immunogenic cell death (ICD), including surface exposure of calreticulin (CRT), the heat shock proteins HSP70 and HSP90, the release of high-mobility group box protein 1 (HMGB1) and the release of ATP from dying cells. We investigated the potential of high hydrostatic pressure (HHP) to induce ICD in human tumor cells. HHP induced the rapid expression of HSP70, HSP90 and CRT on the cell surface. HHP also induced the release of HMGB1 and ATP. The interaction of dendritic cells (DCs) with HHP-treated tumor cells led to a more rapid rate of DC phagocytosis, upregulation of CD83, CD86 and HLA-DR and the release of interleukin IL-6, IL-12p70 and TNF-α. DCs pulsed with tumor cells killed by HHP induced high numbers of tumor-specific T cells. DCs pulsed with HHP-treated tumor cells also induced the lowest number of regulatory T cells. In addition, we found that the key features of the endoplasmic reticulum stress-mediated apoptotic pathway, such as reactive oxygen species production, phosphorylation of the translation initiation factor eIF2α and activation of caspase-8, were activated by HHP treatment. Therefore, HHP acts as a reliable and potent inducer of ICD in human tumor cells.
Subject(s)
Apoptosis/immunology , Dendritic Cells/immunology , Neoplasms/immunology , T-Lymphocytes, Regulatory/immunology , Adenosine Triphosphate/metabolism , Antigens, CD/biosynthesis , B7-2 Antigen/biosynthesis , Calreticulin/biosynthesis , Calreticulin/immunology , Caspase 8/metabolism , Cell Line, Tumor , Endoplasmic Reticulum Stress/immunology , Enzyme Activation/immunology , Eukaryotic Initiation Factor-2/metabolism , HLA-DR Antigens/biosynthesis , HMGB1 Protein/immunology , HMGB1 Protein/metabolism , HSP70 Heat-Shock Proteins/biosynthesis , HSP70 Heat-Shock Proteins/immunology , HSP90 Heat-Shock Proteins/biosynthesis , HSP90 Heat-Shock Proteins/immunology , Humans , Hydrostatic Pressure , Immunoglobulins/biosynthesis , Interleukin-12/metabolism , Interleukin-6/metabolism , Membrane Glycoproteins/biosynthesis , Membrane Proteins/biosynthesis , Phagocytosis/immunology , Phosphorylation , Reactive Oxygen Species/metabolism , Tumor Necrosis Factor-alpha/metabolism , CD83 AntigenABSTRACT
Epithelial ovarian cancer (EOC) is among the top five causes of cancer-related death in women, largely reflecting early, prediagnosis dissemination of malignant cells to the peritoneum. Despite improvements in medical therapies, particularly with the implementation of novel drugs targeting homologous recombination deficiency, the survival rates of patients with EOC remain low. Unlike other neoplasms, EOC remains relatively insensitive to immune checkpoint inhibitors, which is correlated with a tumor microenvironment (TME) characterized by poor infiltration by immune cells and active immunosuppression dominated by immune components with tumor-promoting properties, especially tumor-associated macrophages (TAMs). In recent years, TAMs have attracted interest as potential therapeutic targets by seeking to reverse the immunosuppression in the TME and enhance the clinical efficacy of immunotherapy. Here, we review the key biological features of TAMs that affect tumor progression and their relevance as potential targets for treating EOC. We especially focus on the therapies that might modulate the recruitment, polarization, survival, and functional properties of TAMs in the TME of EOC that can be harnessed to develop superior combinatorial regimens with immunotherapy for the clinical care of patients with EOC.
Subject(s)
Carcinoma , Ovarian Neoplasms , Humans , Female , Carcinoma, Ovarian Epithelial/therapy , Carcinoma, Ovarian Epithelial/pathology , Tumor-Associated Macrophages/pathology , Macrophages , Immunotherapy , Carcinoma/pathology , Tumor MicroenvironmentABSTRACT
While type I interferon (IFN) is best known for its key role against viral infection, accumulating preclinical and clinical data indicate that robust type I IFN production in the tumor microenvironment promotes cancer immunosurveillance and contributes to the efficacy of various antineoplastic agents, notably immunogenic cell death inducers. Here, we report that malignant blasts from patients with acute myeloid leukemia (AML) release type I IFN via a Toll-like receptor 3 (TLR3)-dependent mechanism that is not driven by treatment. While in these patients the ability of type I IFN to stimulate anticancer immune responses was abolished by immunosuppressive mechanisms elicited by malignant blasts, type I IFN turned out to exert direct cytostatic, cytotoxic and chemosensitizing activity in primary AML blasts, leukemic stem cells from AML patients and AML xenograft models. Finally, a genetic signature of type I IFN signaling was found to have independent prognostic value on relapse-free survival and overall survival in a cohort of 132 AML patients. These findings delineate a clinically relevant, therapeutically actionable and prognostically informative mechanism through which type I IFN mediates beneficial effects in patients with AML.
Subject(s)
Antineoplastic Agents , Interferon Type I , Leukemia, Myeloid, Acute , Humans , Leukemia, Myeloid, Acute/pathology , Antineoplastic Agents/therapeutic use , Treatment Outcome , Signal Transduction , Tumor MicroenvironmentABSTRACT
Accumulating evidence indicates that immune checkpoint inhibitors (ICIs) can restore CD8+ cytotoxic T lymphocyte (CTL) functions in preclinical models of acute myeloid leukemia (AML). However, ICIs targeting programmed cell death 1 (PDCD1, best known as PD-1) and cytotoxic T lymphocyte-associated protein 4 (CTLA4) have limited clinical efficacy in patients with AML. Natural killer (NK) cells are central players in AML-targeting immune responses. However, little is known on the relationship between co-inhibitory receptors expressed by NK cells and the ability of the latter to control AML. Here, we show that hepatitis A virus cellular receptor 2 (HAVCR2, best known as TIM-3) is highly expressed by NK cells from AML patients, correlating with improved functional licensing and superior effector functions. Altogether, our data indicate that NK cell frequency as well as TIM-3 expression levels constitute prognostically relevant biomarkers of active immunity against AML.
Subject(s)
Hepatitis A Virus Cellular Receptor 2 , Killer Cells, Natural , Leukemia, Myeloid, Acute , CD8-Positive T-Lymphocytes , Humans , Leukemia, Myeloid, Acute/drug therapy , T-Lymphocytes, CytotoxicABSTRACT
Immunogenic cell death (ICD), a functionally peculiar type of apoptosis, represents a unique way to deliver danger-associated molecular patterns (DAMPs) to the tumor microenvironment. Once emitted by dying cancer cells, DAMPs orchestrate antigen-specific immune responses by acting on both innate and adaptive components of the immune system. Accumulating preclinical and clinical evidence indicates that one of these DAMPs, calreticulin (CALR) represents a novel powerful prognostic biomarker, reflecting the activation of a clinically relevant anticancer immune response in different cancer malignancies. Therefore, the assessment of CALR emission can provide a therapeutic tool for the stratification of cancer patients and the identification of individuals that are intrinsically capable to respond to a particular treatment. Here we describe methods for the quantification of CALR exposure in the tumor microenvironment of cancer patients by flow cytometry and immunohistochemistry.
Subject(s)
Calreticulin/immunology , Flow Cytometry/methods , Immunogenic Cell Death , Immunohistochemistry/methods , Neoplasms/immunology , Animals , Biomarkers, Tumor/analysis , Biomarkers, Tumor/immunology , Calreticulin/analysis , Humans , Neoplasms/pathology , Tumor MicroenvironmentABSTRACT
Natural killer (NK) cells constitute the predominant innate lymphocyte subset that mediates the anti-viral and anti-tumor immune responses. NK cells use an array of innate receptors to sense their environment and to respond to infections, cellular stress and transformation. The resulting NK cell activation, including cytotoxicity and cytokine production, is a fundamental component of the early immune response. The most recent discoveries in NK cell biology have stimulated the translational research that has led to remarkable results for the treatment of human malignancies. Therefore, the rapid isolation of NK cells from the peripheral blood or tumor microenvironment and the subsequent assessment of cytolytic function are crucial to the study of their potency and NK cell-mediated immunosurveillance. Here, we provide protocols for NK cell isolation and the assessment of NK cell cytotoxicity using flow cytometry.
Subject(s)
Cytotoxicity Tests, Immunologic/methods , Cytotoxicity, Immunologic , Flow Cytometry/methods , Killer Cells, Natural/immunology , Cell Separation/methods , Humans , Lymphocyte ActivationABSTRACT
BACKGROUND: The immunological microenvironment of primary high-grade serous carcinomas (HGSCs) has a major impact on disease outcome. Conversely, little is known on the microenvironment of metastatic HGSCs and its potential influence on patient survival. Here, we explore the clinical relevance of the immunological configuration of HGSC metastases. METHODS: RNA sequencing was employed on 24 paired primary tumor microenvironment (P-TME) and metastatic tumor microenvironment (M-TME) chemotherapy-naive HGSC samples. Immunohistochemistry was used to evaluate infiltration by CD8+ T cells, CD20+ B cells, DC-LAMP+ (lysosomal-associated membrane protein 3) dendritic cells (DCs), NKp46+ (natural killer) cells and CD68+CD163+ M2-like tumor-associated macrophages (TAMs), abundance of PD-1+ (programmed cell death 1), LAG-3+ (lymphocyte-activating gene 3) cells, and PD-L1 (programmed death ligand 1) expression in 80 samples. Flow cytometry was used for functional assessments on freshly resected HGSC samples. RESULTS: 1468 genes were differentially expressed in the P-TME versus M-TME of HGSCs, the latter displaying signatures of extracellular matrix remodeling and immune infiltration. M-TME infiltration by immune effector cells had little impact on patient survival. Accordingly, M-TME-infiltrating T cells were functionally impaired, but not upon checkpoint activation. Conversely, cytokine signaling in favor of M2-like TAMs activity appeared to underlie inhibited immunity in the M-TME and poor disease outcome. CONCLUSIONS: Immunosuppressive M2-like TAM infiltrating metastatic sites limit clinically relevant immune responses against HGSCs.
Subject(s)
Biomarkers, Tumor/metabolism , Immunosuppression Therapy/methods , Macrophages/immunology , Ovarian Neoplasms/immunology , Aged , Aged, 80 and over , Female , Humans , Middle Aged , Neoplasm Metastasis , Retrospective Studies , Tumor MicroenvironmentABSTRACT
BACKGROUND: Adjuvanticity, which is the ability of neoplastic cells to deliver danger signals, is critical for the host immune system to mount spontaneous and therapy-driven anticancer immune responses. One of such signals, i.e., the exposure of calreticulin (CALR) on the membrane of malignant cells experiencing endoplasmic reticulum (ER) stress, is well known for its role in the activation of immune responses to dying cancer cells. However, the potential impact of CALR on the immune contexture of primary and metastatic high-grade serous carcinomas (HGSCs) and its prognostic value for patients with HGSC remains unclear. METHOD: We harnessed a retrospective cohort of primary (no = 152) and metastatic (no = 74) tumor samples from HGSC patients to investigate the CALR expression in relation with prognosis and function orientation of the tumor microenvironment. IHC data were complemented with transcriptomic and functional studies on second prospective cohort of freshly resected HGSC samples. In silico analysis of publicly available RNA expression data from 302 HGSC samples was used as a confirmatory approach. RESULTS: We demonstrate that CALR exposure on the surface of primary and metastatic HGSC cells is driven by a chemotherapy-independent ER stress response and culminates with the establishment of a local immune contexture characterized by TH1 polarization and cytotoxic activity that enables superior clinical benefits. CONCLUSIONS: Our data indicate that CALR levels in primary and metastatic HGSC samples have robust prognostic value linked to the activation of clinically-relevant innate and adaptive anticancer immune responses.
Subject(s)
Calreticulin/immunology , Ovarian Neoplasms/immunology , Adult , Aged , Aged, 80 and over , Endoplasmic Reticulum Stress , Female , Humans , Middle Aged , Ovarian Neoplasms/genetics , Prognosis , RNA-Seq , Tumor Microenvironment/genetics , Tumor Microenvironment/immunologyABSTRACT
Increasing evidence suggests that antibody-drug conjugates (ADCs) can enhance anti-tumor immunity and improve clinical outcome. Here, we elucidate the therapeutic efficacy and immune-mediated mechanisms of a novel HER2-targeting ADC bearing a potent anthracycline derivate as payload (T-PNU) in a human HER2-expressing syngeneic breast cancer model resistant to trastuzumab and ado-trastuzumab emtansine. Mechanistically, the anthracycline component of the novel ADC induced immunogenic cell death leading to exposure and secretion of danger-associated molecular signals. RNA sequencing derived immunogenomic signatures and TCRß clonotype analysis of tumor-infiltrating lymphocytes revealed a prominent role of the adaptive immune system in the regulation of T-PNU mediated anti-cancer activity. Depletion of CD8 T cells severely reduced T-PNU efficacy, thus confirming the role of cytotoxic T cells as drivers of the T-PNU mediated anti-tumor immune response. Furthermore, T-PNU therapy promoted immunological memory formation in tumor-bearing animals protecting those from tumor rechallenge. Finally, the combination of T-PNU and checkpoint inhibition, such as α-PD1, significantly enhanced tumor eradication following the treatment. In summary, a novel PNU-armed, HER2-targeting ADC elicited long-lasting immune protection in a murine orthotopic breast cancer model resistant to other HER2-directed therapies. Our findings delineate the therapeutic potential of this novel ADC payload and support its clinical development for breast cancer patients and potentially other HER2 expressing malignancies.
Subject(s)
Anthracyclines/therapeutic use , Antineoplastic Agents, Immunological/therapeutic use , Immunoconjugates/therapeutic use , Mammary Neoplasms, Experimental/drug therapy , Programmed Cell Death 1 Receptor/antagonists & inhibitors , Receptor, ErbB-2/antagonists & inhibitors , Animals , CD8-Positive T-Lymphocytes/drug effects , CD8-Positive T-Lymphocytes/immunology , Cell Line, Tumor , Female , Humans , Immunologic Memory/drug effects , Mammary Neoplasms, Experimental/immunology , Mice, Inbred BALB C , Receptor, ErbB-2/genetics , Trastuzumab/therapeutic useABSTRACT
PURPOSE: In multiple oncological settings, expression of the coinhibitory ligand PD-L1 by malignant cells and tumor infiltration by immune cells expressing coinhibitory receptors such as PD-1, CTLA4, LAG-3, or TIM-3 conveys prognostic or predictive information. Conversely, the impact of these features of the tumor microenvironment on disease outcome among high-grade serous carcinoma (HGSC) patients remains controversial. EXPERIMENTAL DESIGN: We harnessed a retrospective cohort of 80 chemotherapy-naïve HGSC patients to investigate PD-L1 expression and tumor infiltration by CD8+ T cells, CD20+ B cells, DC-LAMP+ dendritic cells as well as by PD-1+, CTLA4+, LAG-3+, and TIM-3+ cells in relation with prognosis and function orientation of the tumor microenvironment. IHC data were complemented with transcriptomic and functional studies on a second prospective cohort of freshly resected HGSC samples. In silico analysis of publicly available RNA expression data from 308 HGSC samples was used as a confirmatory approach. RESULTS: High levels of PD-L1 and high densities of PD-1+ cells in the microenvironment of HGSCs were strongly associated with an immune contexture characterized by a robust TH1 polarization and cytotoxic orientation that enabled superior clinical benefits. Moreover, PD-1+TIM-3+CD8+ T cells presented all features of functional exhaustion and correlated with poor disease outcome. However, although PD-L1 levels and tumor infiltration by TIM-3+ cells improved patient stratification based on the intratumoral abundance of CD8+ T cells, the amount of PD-1+ cells failed to do so. CONCLUSIONS: Our data indicate that PD-L1 and TIM-3 constitute prognostically relevant biomarkers of active and suppressed immune responses against HGSC, respectively.
Subject(s)
Biomarkers, Tumor/metabolism , CD8-Positive T-Lymphocytes/immunology , Carcinoma, Ovarian Epithelial/immunology , Cystadenocarcinoma, Serous/immunology , Gene Expression Regulation, Neoplastic , Hepatitis A Virus Cellular Receptor 2/metabolism , Lymphocytes, Tumor-Infiltrating/immunology , Adult , Aged , B7-H1 Antigen/immunology , B7-H1 Antigen/metabolism , Biomarkers, Tumor/immunology , CTLA-4 Antigen/immunology , CTLA-4 Antigen/metabolism , Carcinoma, Ovarian Epithelial/metabolism , Carcinoma, Ovarian Epithelial/pathology , Cystadenocarcinoma, Serous/metabolism , Cystadenocarcinoma, Serous/pathology , Female , Hepatitis A Virus Cellular Receptor 2/immunology , Humans , Lysosomal Membrane Proteins/immunology , Lysosomal Membrane Proteins/metabolism , Middle Aged , Neoplasm Proteins/immunology , Neoplasm Proteins/metabolism , Prognosis , Retrospective Studies , Survival RateABSTRACT
The death of cancer cells can be categorized as either immunogenic (ICD) or nonimmunogenic, depending on the initiating stimulus. The immunogenic processes of immunogenic cell death are mainly mediated by damage-associated molecular patterns (DAMPs), which include surface exposure of calreticulin (CRT), secretion of adenosine triphosphate (ATP), release of non-histone chromatin protein high-mobility group box 1 (HMGB1) and the production of type I interferons (IFNs). DAMPs are recognized by various receptors that are expressed by antigen-presenting cells (APCs) and potentiate the presentation of tumor antigens to T lymphocytes. Accumulating evidence indicates that CRT exposure constitutes one of the major checkpoints, that determines the immunogenicity of cell death both in vitro and in vivo in mouse models. Moreover, recent studies have identified CRT expression on tumor cells not only as a marker of ICD and active anti-tumor immune reactions but also as a major predictor of a better prognosis in various cancers. Here, we discuss the recent information on the CRT capacity to activate anticancer immune response as well as its prognostic and predictive role for the clinical outcome in cancer patients.
Subject(s)
Antigen-Presenting Cells/immunology , Calreticulin/metabolism , Neoplasms/metabolism , T-Lymphocytes/immunology , Adenosine Triphosphate/metabolism , Animals , Calreticulin/genetics , Disease Models, Animal , Endoplasmic Reticulum Stress , Gene Expression Regulation, Neoplastic , HMGB1 Protein/genetics , HMGB1 Protein/metabolism , Humans , Immunity , Interferon Type I/metabolism , Mice , Neoplasms/diagnosis , PrognosisABSTRACT
A high density of tumor-infiltrating CD8+ T cells and CD20+ B cells correlates with prolonged survival in patients with a wide variety of human cancers, including high-grade serous ovarian carcinoma (HGSC). However, the potential impact of mature dendritic cells (DCs) in shaping the immune contexture of HGSC, their role in the establishment of T cell-dependent antitumor immunity, and their potential prognostic value for HGSC patients remain unclear. We harnessed immunohistochemical tests and biomolecular analyses to demonstrate that a high density of tumor-infiltrating DC-LAMP+ DCs is robustly associated with an immune contexture characterized by TH1 polarization and cytotoxic activity. We showed that both mature DCs and CD20+ B cells play a critical role in the generation of a clinically-favorable cytotoxic immune response in HGSC microenvironment. In line with this notion, robust tumor infiltration by both DC-LAMP+ DCs and CD20+ B cells was associated with most favorable overall survival in two independent cohorts of chemotherapy-naïve HGSC patients. Our findings suggest that the presence of mature, DC-LAMP+ DCs in the tumor microenvironment may represent a novel, powerful prognostic biomarker for HGSC patients that reflects the activation of clinically-relevant anticancer immunity.
Subject(s)
Carcinoma/immunology , Carcinoma/pathology , Dendritic Cells/immunology , Dendritic Cells/pathology , Ovarian Neoplasms/immunology , Ovarian Neoplasms/pathology , Tumor Microenvironment/immunology , Adult , Aged , Aged, 80 and over , Biomarkers , Carcinoma/mortality , Dendritic Cells/metabolism , Female , Humans , Immunohistochemistry , Immunophenotyping , Kaplan-Meier Estimate , Lymphocytes, Tumor-Infiltrating/immunology , Lymphocytes, Tumor-Infiltrating/metabolism , Lymphocytes, Tumor-Infiltrating/pathology , Middle Aged , Neoplasm Staging , Ovarian Neoplasms/mortality , Ovarian Neoplasms/therapy , Prognosis , Proportional Hazards Models , T-Lymphocyte Subsets/immunology , T-Lymphocyte Subsets/metabolism , T-Lymphocyte Subsets/pathologyABSTRACT
Dendritic cell (DC)-based vaccines pulsed with high hydrostatic pressure (HHP)-inactivated tumor cells have recently been shown to be a promising tool for prostate cancer chemoimmunotherapy. In this study, DC-based vaccines, both pulsed and unpulsed, were as effective as docetaxel (DTX) in reducing prostate tumors in the orthotopic transgenic adenocarcinoma of the mouse prostate (TRAMP) model. However, we did not observe any additive or synergic effects of chemoimmunotherapy on the tumor growth, while only the combination of DTX and pulsed dendritic cells resulted in significantly lower proliferation detected by Ki67 staining in histological samples. The DC-based vaccine pulsed with HHP-treated tumor cells was also combined with another type of cytostatic, cyclophosphamide, with similar results. In another clinically relevant setting, minimal residual tumor disease after surgery, administration of DC-based vaccines after the surgery of poorly immunogenic transplanted TRAMP-C2, as well as in immunogenic TC-1 tumors, reduced the growth of tumor recurrences. To identify the effector cell populations after DC vaccine application, mice were twice immunized with both pulsed and unpulsed DC vaccine, and the cytotoxicity of the spleen cells populations was tested. The effector cell subpopulations were defined as CD4+ and NK1.1+, which suggests rather unspecific therapeutic effects of the DC-based vaccines in our settings. Taken together, our data demonstrate that DC-based vaccines represent a rational tool for the treatment of human prostate cancer.
ABSTRACT
High hydrostatic pressure (HHP) promotes key characteristics of immunogenic cell death (ICD), in thus far resembling immunogenic chemotherapy and ionizing irradiation. Here, we demonstrate that cancer cells succumbing to HHP induce CD4+ and CD8+ T cell-dependent protective immunity in vivo. Moreover, we show that cell death induction by HHP relies on the overproduction of reactive oxygen species (ROS), causing rapid establishment of the integrated stress response, eIF2α phosphorylation by PERK, and sequential caspase-2, -8 and -3 activation. Non-phosphorylatable eIF2α, depletion of PERK, caspase-2 or -8 compromised calreticulin exposure by cancer cells succumbing to HHP but could not inhibit death. Interestingly, the phagocytosis of HHP-treated malignant cells by dendritic cells was suppressed by the knockdown of caspase-2 in the former. Thus, caspase-2 mediates a key function in the interaction between dying cancer cells and antigen presenting cells. Our results indicate that the ROSâPERKâeIF2αâcaspase-2 signaling pathway is central for the perception of HHP-driven cell death as immunogenic.
ABSTRACT
High hydrostatic pressure (HHP) has been shown to induce immunogenic cell death of cancer cells, facilitating their uptake by dendritic cells (DC) and subsequent presentation of tumor antigens. In the present study, we demonstrated immunogenicity of the HHP-treated tumor cells in mice. HHP was able to induce immunogenic cell death of both TC-1 and TRAMP-C2 tumor cells, representing murine models for human papilloma virus-associated tumors and prostate cancer, respectively. HHP-treated cells induced stronger immune responses in mice immunized with these tumor cells, documented by higher spleen cell cytotoxicity and increased IFNγ production as compared to irradiated tumor cells, accompanied by suppression of tumor growth in vivo in the case of TC-1 tumors, but not TRAMP-C2 tumors. Furthermore, HHP-treated cells were used for DC-based vaccine antigen pulsing. DC co-cultured with HHP-treated tumor cells and matured by a TLR 9 agonist exhibited higher cell surface expression of maturation markers and production of IL-12 and other cytokines, as compared to the DC pulsed with irradiated tumor cells. Immunization with DC cell-based vaccines pulsed with HHP-treated tumor cells induced high immune responses, detected by increased spleen cell cytotoxicity and elevated IFNγ production. The DC-based vaccine pulsed with HHP-treated tumor cells combined with docetaxel chemotherapy significantly inhibited growth of both TC-1 and TRAMP-C2 tumors. Our results indicate that DC-based vaccines pulsed with HHP-inactivated tumor cells can be a suitable tool for chemoimmunotherapy, particularly with regard to the findings that poorly immunogenic TRAMP-C2 tumors were susceptible to this treatment modality.