The sentinel within: exploiting the immune system for cancer biomarkers

The release of proteins from tumors triggers an immune response in cancer patients. These tumor antigens arise from several mechanisms including tumor-specific alterations in protein expression, mutation, folding, degradation, or intracellular localization. Responses to most tumor antigens are rarely observed in healthy individuals, making the response itself a biomarker that betrays the presence of underlying cancer. Antibody immune responses show promise as clinical biomarkers because antibodies have long half-lives in serum, are easy to measure, and are stable in blood samples. However, our understanding of the specificity and the impact of the immune response in early stages of cancer is limited. The immune response to cancer, whether endogenous or driven by vaccines, involves highly specific T lymphocytes (which target tumor-derived peptides bound to self-MHC proteins) and B lymphocytes (which generate antibodies to tumor-derived proteins). T cell target antigens have been identified either by expression cloning from tumor cDNA libraries, or by prediction based on patterns of antigen expression ("reverse immunology"). B cell targets have been similarly identified using the antibodies in patient sera to screen cDNA libraries derived from tumor cell lines. This review focuses on the application of recent advances in proteomics for the identification of tumor antigens. These advances are opening the door for targeted vaccine development, and for using immune response signatures as biomarkers for cancer diagnosis and monitoring.     

CD47分子与抗肿瘤免疫

肿瘤进展与人免疫系统间的联系已经被广泛研究,有许多免疫分子已被证实参与其中。CD47（整合素相关蛋白）为一种免疫球蛋白超家族成员,在人免疫系统中发挥着重要功能。研究表明CD47在肿瘤细胞表面也有高表达,其高表达与肿瘤的生长、转移及复发等密切相关。肿瘤细胞表面的CD47与巨噬细胞表面的SIRPα相互作用,并发出“别吃我”的免疫抑制性信号,从而保护肿瘤细胞免受巨噬细胞吞噬。因此,开发以CD47为靶点的拮抗剂可阻断此抑制性信号,从而增强巨噬细胞的吞噬效应,以达到增强抗肿瘤免疫反应的目的。最新研究证实,CD47拮抗剂在T细胞介导的抗肿瘤免疫反应中也发挥了重要作用。本文将对CD47分子的结构功能、在抗肿瘤免疫反应中的作用及以其为靶点的拮抗剂研究进展进行综述,以期为进一步的药物开发及临床研究等提供参考。     

Striking immunodominance hierarchy of naturally occurring CD8+ and CD4+ T cell responses to tumor antigen NY-ESO-1

Immunodominance has been well-demonstrated in many antiviral and antibacterial systems, but much less so in the setting of immune responses against cancer. Tumor Ag-specific CD8+ T cells keep cancer cells in check via immunosurveillance and shape tumor development through immunoediting. Because most tumor Ags are self Ags, the breadth and depth of antitumor immune responses have not been well-appreciated. To design and develop antitumor vaccines, it is important to understand the immunodominance hierarchy and its underlying mechanisms, and to identify the most immunodominant tumor Ag-specific T cells. We have comprehensively analyzed spontaneous cellular immune responses of one individual and show that multiple tumor Ags are targeted by the patient's immune system, especially the "cancer-testis" tumor Ag NY-ESO-1. The pattern of anti-NY-ESO-1 T cell responses in this patient closely resembles the classical broad yet hierarchical antiviral immunity and was confirmed in a second subject.     

TXLNA enhances TBK1 phosphorylation by suppressing PPM1B recruitment

In recent years, there has been a notable increase in cancer incidence and mortality, and immune abnormalities have been closely linked to malignancy development. TANK-binding kinase 1 (TBK1) is a non-classical IκB kinase that regulates interferon and NF-κB signaling pathways and plays a crucial role in innate immunity. Recent studies have shown high expression levels of TBK1 and increased activity in various tumor cells, suggesting its involvement in the development and progression of multiple cancers. Targeting TBK1 for tumor therapy may be a possibility. However, little is known about the abnormal activation and dynamic regulation of TBK1 in cancer. First, we utilized the BioID biotinylation technique combined with TMT-based quantitative proteomics to analyze the TBK1 interacting proteins. Our results revealed that TXLNA interacts with TBK1 and binds to the α-helical scaffold of TBK1. The expression of TXLNA could affect the S172 phosphorylation of TBK1. PPM1B is a phosphatase that can dephosphorylate TBK1 S172, so we used the APEX2 proximity labeling technique combined with TMT-based quantitative proteomics to explore the interacting proteins of PPM1B and search for the regulatory pathway of TXLNA on TBK1 phosphorylation. We found that PPM1B interacts with TXLNA. Based on these results, we further found that TXLNA impairs the binding of PPM1B to TBK1, inhibiting the dephosphorylation of TBK1 and contributing to the abnormal enhancement of TBK1 activity in cancer cells. This study sheds light on the potential mechanism of aberrant activation and dynamic regulation of TBK1 in tumors and provides a potential target for tumor therapy.     

Non‐viral gene delivery for cancer immunotherapy

Recent decades have witnessed the revolutionary development of cancer immunotherapies, which boost cancer‐specific immune responses for long‐term tumor regression. However, immunotherapy still has limitations, including off‐target side effects, long processing times and limited patient responses. These disadvantages of current immunotherapy are being addressed by improving our understanding of the immune system, as well as by establishing combinational approaches. Advanced biomaterials and gene delivery systems overcome some of these delivery issues, harnessing adverse effects and amplifying immunomodulatory effects, and are superior to standard formulations with respect to eliciting antitumor immunity. Nucleic acid‐based nanostructures have diverse functions, ranging from gene expression and gene regulation to pro‐inflammatory effects, as well as the ability to specifically bind different molecules. A brief overview is provided of the recent advances in the non‐viral gene delivery methods that are being used to activate cancer‐specific immune responses. Furthermore, the tumor microenvironment‐responsive synergistic strategies that modulate the immune response by targeting various signaling pathways are discussed. Nanoparticle‐based non‐viral gene delivery strategies have great potential to be implemented in the clinic for cancer immunotherapy.     

Synthetic peptides as cancer vaccines

Effective cancer therapy or prevention has been the dream of physicians and scientists for many years. Although we are still very far from our ultimate goal of cancer prevention, significant milestones have been realized in terms of our knowledge base and understanding of the pathogenesis of cancerous cells and the involvement of the immune system against both self- and virus-associated tumor antigens. Immunotherapeutic strategies are now accepted to being superior in terms of the exquisite specificity that they offer in targeting only tumor cells as opposed to the existent chemotherapy or radiation therapy that is more general and invasive with many associated side effects. There are several immunotherapeutic strategies that are currently under investigation. This review primarily focuses on the significant advances made in the use of synthetic peptides in the development of subunit cancer vaccines. We have attempted to highlight some of the fundamental issues regarding antigen processing and presentation, Major Histocompatibility Complex (MHC) restriction, T-cell help, structural determinants in antibody recognition, and the use of these concepts in the rational design and delivery of peptide vaccines to elicit protective humoral and cell mediated immune responses. The recent use of costimulatory molecules and cytokines to augment immune responses also has been discussed along with the contributions of our laboratory to the field of synthetic peptide vaccine development.     

Isolation of immune cells from primary tumors

Tumors create a unique immunosuppressive microenvironment (tumor microenvironment, TME) whereby leukocytes are recruited into the tumor by various chemokines and growth factors. However, once in the TME, these cells lose the ability to promote anti-tumor immunity and begin to support tumor growth and down-regulate anti-tumor immune responses. Studies on tumor-associated leukocytes have mainly focused on cells isolated from tumor-draining lymph nodes or spleen due to the inherent difficulties in obtaining sufficient cell numbers and purity from the primary tumor. While identifying the mechanisms of cell activation and trafficking through the lymphatic system of tumor bearing mice is important and may give insight to the kinetics of immune responses to cancer, in our experience, many leukocytes, including dendritic cells (DCs), in tumor-draining lymph nodes have a different phenotype than those that infiltrate tumors. Furthermore, we have previously demonstrated that adoptively-transferred T cells isolated from the tumor-draining lymph nodes are not tolerized and are capable of responding to secondary stimulation in vitro unlike T cells isolated from the TME, which are tolerized and incapable of proliferation or cytokine production. Interestingly, we have shown that changing the tumor microenvironment, such as providing CD4(+) T helper cells via adoptive transfer, promotes CD8(+) T cells to maintain pro-inflammatory effector functions. The results from each of the previously mentioned studies demonstrate the importance of measuring cellular responses from TME-infiltrating immune cells as opposed to cells that remain in the periphery. To study the function of immune cells which infiltrate tumors using the Miltenyi Biotech isolation system, we have modified and optimized this antibody-based isolation procedure to obtain highly enriched populations of antigen presenting cells and tumor antigen-specific cytotoxic T lymphocytes. The protocol includes a detailed dissection of murine prostate tissue from a spontaneous prostate tumor model (TRansgenic Adenocarcinoma of the Mouse Prostate -TRAMP) and a subcutaneous melanoma (B16) tumor model followed by subsequent purification of various leukocyte populations.     

Dissecting tumor responsiveness to immunotherapy: the experience of peptide-based melanoma vaccines

Recent years have witnessed important breakthroughs in our understanding of tumor immunology. A variety of immunotherapeutic strategies has shown that immune manipulation can induce the regression of established cancer in humans. The identification of the genes encoding tumor-associated antigens (TAA) and the development of means for immunizing against these antigens have opened new avenues for the development of an effective anticancer immunotherapy. However, an efficient immune response against tumor requires an intricate cross-talk between cancer and immune system cells, which is still poorly understood. Only when the molecular basis underlying tumor susceptibility to an immune response is deciphered could new therapeutic strategies be designed to fit biologically defined mechanisms of cancer immune rejection. In this article, we address some of the critical issues that have been identified in cancer immunotherapy, in part from our own studies on immune therapies in melanoma patients treated with peptide-based vaccination regimens. This is not meant to be a comprehensive overview of the immunological phenomena accompanying cancer patient vaccination but rather emphasizes some emergent findings, puzzling controversies and unanswered questions that characterize this complex field of oncology. In addition to reviewing the main immunological concepts underlying peptide-based vaccination, we also review the available data regarding naturally occurring and therapeutically induced anticancer immune response, both at the peripheral and intratumoral level. The hypothesized role of innate immunity in predetermining tumor responsiveness to immunotherapeutic manipulation is also discussed.     

Spontaneous immune responses to sporadic tumors: tumor-promoting, tumor-protective or both?

Cancer cells cannot develop into invasive cancers without interactions with cells and soluble mediators present in the tumor microenvironment. Accumulating evidence indicates that the immune system is a critical determinant of malignant outgrowth; however, the tumor-modulating effects of spontaneous immune responses towards nascent malignancies are rather paradoxical. Both cancer-protective and cancer-promoting features of the immune system have been described. This review will discuss the role of the dynamic inflammatory tumor microenvironment during cancer development and progression, and will focus on the intriguing question: “Do malignancies develop in spite of—or because of—spontaneous immune responses?” Special emphasis will be put on recent progress in our understanding of the immune system’s double-edged sword function during de novo carcinogenesis.     

Impacts and mechanisms of metabolic reprogramming of tumor microenvironment for immunotherapy in gastric cancer

Metabolic disorders and abnormal immune function changes occur in tumor tissues and cells to varying degrees. There is increasing evidence that reprogrammed energy metabolism contributes to the development of tumor suppressive immune microenvironment and influences the course of gastric cancer (GC). Current studies have found that tumor microenvironment (TME) also has important clinicopathological significance in predicting prognosis and therapeutic efficacy. Novel approaches targeting TME therapy, such as immune checkpoint blockade (ICB), metabolic inhibitors and key enzymes of immune metabolism, have been involved in the treatment of GC. However, the interaction between GC cells metabolism and immune metabolism and how to make better use of these immunotherapy methods in the complex TME in GC are still being explored. Here, we discuss how metabolic reprogramming of GC cells and immune cells involved in GC immune responses modulate anti-tumor immune responses, as well as the effects of gastrointestinal flora in TME and GC. It is also proposed how to enhance anti-tumor immune response by understanding the targeted metabolism of these metabolic reprogramming to provide direction for the treatment and prognosis of GC.Subject terms: Cancer, Mechanisms of disease     

Tumor vaccine: current trends in antigen specific immunotherapy

Effective cancer treatment to prevent the tumor growth as well as to stop its recurrence is the dream of oncologists. Currently available therapeutic measures like, radiotherapy and chemotherapy, often suffer from severe toxicity and lack of specificity of the drug towards tumor cells. Another promising approach is the 'immunotherapy', in which either the immune system is activated by tumor vaccine to combat the tumor growth or antitumor antibodies can be used. Vaccination can stimulate humoral, cellular and innate immune systems to generate various effector molecules, like antibody, cytotoxic T cells, cytokines etc. In antigen specific immunotherapy, the immune system can be stimulated actively by antigen based tumor vaccine to kill only those tumor cells, having expression of the particular tumor associated antigen. Different experimental, preclinical and clinical studies have proved that generated immune responses are effective to restrict the tumor growth. Useful strategies of antigen specific immunotherapy and outcome of various laboratory and clinic based studies are discussed.     

Interleukin 18: Friend or foe in cancer

In the last few years, the field of tumor immunology has significantly expanded and its boundaries, never particularly clear, have become less distinct. Although the immune system plays an important role in controlling tumor growth, it has also become clear that tumor growth can be promoted by inflammatory immune responses. A good example that exemplifies the ambiguous role of the immune system in cancer progression is represented by interleukin 18 (IL-18) that was first identified as an interferon-γ-inducing factor (IGIF) involved in T helper type-1 (Th1) immune response. The expression and secretion of IL-18 have been observed in various cell types from immune cells to circulating cancer cells. In this review we highlighted the multiple roles played by IL-18 in immune regulation, cancer progression and angiogenesis and the clinical potential that may result from such understanding.     

Dual roles of immune cells and their factors in cancer development and progression

Traditional wisdom holds that intact immune responses, such as immune surveillance or immunoediting, are required for preventing and inhibiting tumor development; but recent evidence has also indicated that unresolved immune responses, such as chronic inflammation, can promote the growth and progression of cancer. Within the immune system, cytotoxic CD8(+) and CD4(+) Th1 T cells, along with their characteristically produced cytokine IFN-γ, function as the major anti-tumor immune effector cells, whereas tumor associated macrophages (TAM) or myeloid-derived suppressive cells (MDSC) and their derived cytokines IL-6, TNF, IL-1β and IL-23 are generally recognized as dominant tumor-promoting forces. However, the roles played by Th17 cells, CD4(+) CD25(+) Foxp3(+) regulatory T lymphocytes and immunoregulatory cytokines such as TGF-β in tumor development and survival remain elusive. These immune cells and the cellular factors produced from them, including both immunosuppressive and inflammatory cytokines, play dual roles in promoting or discouraging cancer development, and their ultimate role in cancer progression may rely heavily on the tumor microenvironment and the events leading to initial propagation of carcinogenesis.     

GSDMs家族蛋白介导细胞焦亡在抗肿瘤免疫中的作用

细胞焦亡是一种调节性细胞死亡方式。Gasdermine（GSDMs）是一类执行细胞焦亡的胞内蛋白质。虽然GSDMs表达后的完整蛋白质不具有活性,但能被某些蛋白水解酶激活。被激活的GSDMs N端在质膜上穿孔,导致细胞裂解,引起细胞内的促炎分子及损伤相关分子模式（danger-associated molecular patterns,DAMPs）迅速有效地从焦亡细胞中释放,从而引发炎症和免疫反应。焦亡细胞促进抗肿瘤免疫作用可能涉及细胞毒性T淋巴细胞对肿瘤细胞的杀伤。本文介绍GSDMs介导的细胞焦亡及细胞焦亡过程中引发促炎症和免疫反应的关键分子,并且探讨细胞焦亡对肿瘤治疗的有利及不利因素,以期更好地了解细胞焦亡对肿瘤免疫微环境的影响及对肿瘤免疫治疗的作用,有助于促进恶性肿瘤治疗策略的改进。     

Interferon-alpha in tumor immunity and immunotherapy

Interferon-alpha (IFN-alpha) is a pleiotropic cytokine belonging to type I IFN, currently used in cancer patients. Early studies in mouse tumor models have shown the importance of host immune mechanisms in the generation of a long-lasting antitumor response to type I IFN. Recent studies have underscored new immunomodulatory effects of IFN-alpha, including activities on T and dendritic cells, which may explain IFN-induced tumor immunity. Reports on new immune correlates in cancer patients responding to IFN-alpha represent additional evidence on the importance of the interactions of IFN-alpha with the immune system for the generation of durable antitumor response. This knowledge, together with results from studies on genetically modified tumor cells expressing IFN-alpha, suggest novel strategies for using these cytokines in cancer immunotherapy and in particular the use of IFN-alpha as an immune adjuvant for the development of cancer vaccines.     

Thyroid tumors: novel insights from proteomic studies

Several genomics-based techniques have been applied in the last decade to the molecular characterization of cancer, which has led to a variety of applications suitable for improved diagnosis, prognosis and prediction of outcome to treatment. Proteomics-based approaches have also been seen as crucial to the discovery of biomarkers for early diagnosis and prognosis of tumors, as well as for a better understanding of the molecular bases of cancer. Accordingly, proteomic techniques have been used extensively for a better molecular characterization of thyroid tumors. In this field, three main directions have been preceded: first, proteomic studies of model systems; second, proteomics of thyroid tumor specimens; and third, serum proteomics. In this review, we describe the most relevant results that have been obtained for tumors derived from thyroid follicular cells using various proteomic approaches.     

Developing oncolytic viruses for clinical use: A consortium approach

The use of oncolytic viruses forms an appealing approach for cancer treatment. On the one hand the viruses replicate in, and kill, tumor cells, leading to their intra-tumoral amplification. On the other hand the viral infection will activate virus-directed immune responses, and may trigger immune responses directed against tumor cells and tumor antigens. To date, a wide variety of oncolytic viruses is being developed for use in cancer treatment. While the development of oncolytic viruses has often been initiated by researchers in academia and other public institutions, a large majority of the final product development and the testing of these products in clinical trials is industry led. As a consequence relatively few pre-clinical and clinical studies evaluated different oncolytic viruses in competitive side-by-side preclinical or clinical studies. In this review we will summarize the steps and considerations essential in the development and characterization of oncolytic viruses, and describe our multidisciplinary academic consortium, which involves a dozen departments in three different Dutch universities, collaborating in the development of oncolytic viruses. This consortium has the ambition to develop a small series of oncolytic viruses and to evaluate these in various cancers.     

Combinational adenovirus-mediated gene therapy and dendritic cell vaccine in combating well-established tumors

Recent developments in tumor immunology and biotechnology have made cancer gene therapy and immunotherapy feasible. The current efforts for cancer gene therapy mainly focus on using immunogenes, chemogenes and tumor suppressor genes. Central to all these therapies is the development of efficient vectors for gene therapy. By far, adenovirus （AdV）-mediated gene therapy is one of the most promising approaches, as has confirmed by studies relating to animal tumor models and clinical trials. Dendritic cells （DCs） are highly efficient, specialized antigen-presenting cells, and DC- based tumor vaccines are regarded as having much potential in cancer immunotherapy. Vaccination with DCs pulsed with tumor peptides, lysates, or RNA, or loaded with apoptotic/necrotic tumor cells, or engineered to express certain cytokines or chemokines could induce significant antitumor cytotoxic T lymphocyte （CTL） responses and antitumor immunity. Although both AdV-mediated gene therapy and DC vaccine can both stimulate antitumor immune responses, their therapeutic efficiency has been limited to generation of prophylactic antitumor immunity against re-challenge with the parental tumor cells or to growth inhibition of small tumors. However, this approach has been unsuccessful in combating well-established tumors in animal models. Therefore, a major strategic goal of current cancer immunotherapy has become the development of novel therapeutic strategies that can combat well-established tumors, thus resembling real clinical practice since a good proportion of cancer patients generally present with significant disease. In this paper, we review the recent progress in AdV-mediated cancer gene therapy and DC-based cancer vaccines, and discuss combined immunotherapy including gene therapy and DC vaccines. We underscore the fact that combined therapy may have some advantages in combating well-established tumors vis-a-vis either modality administered as a monotherapy.