The redox protein HMGB1 regulates cell death and survival in cancer treatment

Metabolic and therapeutic stress activates several signal transduction pathways and releases damageassociated molecular pattern molecules (DAMPs) that regulate cell death and cell survival. The prototypical DAMP, high-mobility group box 1 protein (HMGB1) is released with sustained autophagy, late apoptosis and necrosis. Our recent findings reveal that the HMGB1 protein triggers autophagy or apoptosis in cancer cells, depending on its redox status. Reducible HMGB1 binds to the receptor for advanced glycation end products (RAGE), induces Beclin 1-dependent autophagy and promotes pancreatic or colon tumor cell line resistance to chemotherapeutic agents or ionizing radiation. In contrast, oxidized HMGB1 increases the cytotoxicity of these agents and induces apoptosis via the mitochondrial pathway. This suggests a new function for HMGB1 within the tumor microenvironment, regulating cell death and survival and suggests that it plays an important functional role in cross-regulating apoptosis and autophagy.     

Many faces of DAMPs in cancer therapy

A new concept of immunogenic cell death (ICD) has recently been proposed. The immunogenic characteristics of this cell death mode are mediated mainly by molecules called ‘damage-associated molecular patterns'' (DAMPs), most of which are recognized by pattern recognition receptors. Some DAMPs are actively emitted by cells undergoing ICD (e.g. calreticulin (CRT) and adenosine triphosphate (ATP)), whereas others are emitted passively (e.g. high-mobility group box 1 protein (HMGB1)). Recent studies have demonstrated that these DAMPs play a beneficial role in anti-cancer therapy by interacting with the immune system. The molecular pathways involved in translocation of CRT to the cell surface and secretion of ATP from tumor cells undergoing ICD are being elucidated. However, it has also been shown that the same DAMPs could contribute to progression of cancer and promote resistance to anticancer treatments. In this review, we will critically evaluate the beneficial and detrimental roles of DAMPs in cancer therapy, focusing mainly on CRT, ATP and HMGB1.     

Danger signalling during cancer cell death: origins,plasticity and regulation

Accumulating data indicates that following anti-cancer treatments, cancer cell death can be perceived as immunogenic or tolerogenic by the immune system. The former is made possible due to the ability of certain anti-cancer modalities to induce immunogenic cell death (ICD) that is associated with the emission of damage-associated molecular patterns (DAMPs), which assist in unlocking a sequence of events leading to the development of anti-tumour immunity. In response to ICD inducers, activation of endoplasmic reticulum (ER) stress has been identified to be indispensable to confer the immunogenic character of cancer cell death, due to its ability to coordinate the danger signalling pathways responsible for the trafficking of vital DAMPs and subsequent anti-cancer immune responses. However, in recent times, certain processes apart from ER stress have emerged (e.g., autophagy and possibly viral response-like signature), which have the ability to influence danger signalling. In this review, we discuss the molecular nature, emerging plasticity in the danger signalling mechanisms and immunological impact of known DAMPs in the context of immunogenic cancer cell death. We also discuss key effector mechanisms modulating the interface between dying cancer cells and the immune cells, which we believe are crucial for the therapeutic relevance of ICD in the context of human cancers, and also discuss the influence of experimental conditions and animal models on these.     

Capsaicin as an inducer of damage-associated molecular patterns (DAMPs) of immunogenic cell death (ICD) in human bladder cancer cells

Few conventional cytotoxic anticancer therapeutics induce immunogenic cell death (ICD). This means that they induce tumor cells to undergo apoptosis while eliciting the emission of a spatiotemporal-defined combination of damage-associated molecular patterns (DAMPs) decoded by the immune system to activate antitumor immunity effective for long-term therapeutic success. The neurotoxin capsaicin (CPS) can induce both cancer cell apoptosis and immune-mediated tumor regression. In the present study, we investigated whether CPS is capable of eliciting the emission of ICD hallmarks in human bladder cancer cell lines undergoing apoptosis. We demonstrated that CPS induces pre- and early apoptotic cell surface exposure of calreticulin (CRT), HSP90, and HSP70 as well as ATP release. Moreover, CRT exposure was prevented by inhibition of endoplasmic reticulum–Golgi traffic by brefeldin A. Furthermore, high-mobility group box 1, HSP90, and HSP70 were passively released at late apoptotic stages. We provide the first evidence that CPS is an inducer of ICD hallmarks, suggesting CPS as a novel potential immunogenic cytotoxic agent.     

DAMPs and autophagy

Autophagy is a lysosome-mediated catabolic process involving the degradation of intracellular contents (e.g., proteins and organelles) as well as invading microbes (e.g., parasites, bacteria and viruses). Multiple forms of cellular stress can stimulate this pathway, including nutritional imbalances, oxygen deprivation, immunological response, genetic defects, chromosomal anomalies and cytotoxic stress. Damage-associated molecular pattern molecules (DAMPs) are released by stressed cells undergoing autophagy or injury, and act as endogenous danger signals to regulate the subsequent inflammatory and immune response. A complex relationship exists between DAMPs and autophagy in cellular adaption to injury and unscheduled cell death. Since both autophagy and DAMPs are important for pathogenesis of human disease, it is crucial to understand how they interplay to sustain homeostasis in stressful or dangerous environments.     

DAMP-mediated autophagy contributes to drug resistance

Damage-associated molecular pattern molecules (DAMPs) are cellularly derived molecules that can initiate and perpetuate immune responses following trauma, ischemia and other types of tissue damage in the absence of pathogenic infection. High mobility group box 1 (HMGB1) is a prototypical DAMP and is associated with the hallmarks of cancer. Recently we found that HMGB1 release after chemotherapy treatment is a critical regulator of autophagy and a potential drug target for therapeutic interventions in leukemia. Overexpression of HMGB1 by gene transfection rendered leukemia cells resistant to cell death; whereas depletion or inhibition of HMGB1 and autophagy by RNA interference or pharmacological inhibitors increased the sensitivity of leukemia cells to chemotherapeutic drugs. HMGB1 release sustains autophagy as assessed by microtubule-associated protein 1 light chain 3 (LC3) lipidation, redistribution of LC3 into cytoplasmic puncta, degradation of p62 and accumulation of autophagosomes and autolysosomes. Moreover, these data suggest a role for HMGB1 in the regulation of autophagy through the PI3KC3-MEKERK: pathway, supporting the notion that HMGB1-induced autophagy promotes tumor resistance to chemotherapy.     

Blocking the interleukin 2 (IL2)-induced systemic autophagic syndrome promotes profound antitumor effects and limits toxicity

Cancer is the leading cause of death in the United States in those dying under the age of 85. Although cancer is increasingly controlled as a chronic disease, true cures of patients with metastatic epithelial malignancies have rarely been obtained with currently available systemic therapies. For example, administration of high-dose recombinant interleukin 2 (IL2), enhancing cytolytic immune cell proliferation and delivery, promotes complete antitumor responses in < 10% of treated individuals. Means to reduce the toxicity, attributed to a cytokine storm and an associated “systemic autophagic syndrome” as well as enhance efficacy and increase the potential set of malignancies in which it is applied (currently patients with renal cancer and melanoma) would be of great interest. IL2 promotes both T-cell and NK cell induction of immune cell-mediated autophagy (iC-MA) in tumor targets. We have demonstrated that HMGB1 is detected at high levels in the serum of IL2-treated mice with translocation to the cytoplasm from the nucleus in the liver, consistent with HMGB1’s release in response to stress, and ability to sustain autophagy. Limiting autophagy in mice with coadministration of chloroquine (CQ) diminishes serum levels of HMGB1, cytokines (IFNG and IL6 but not IL18), and autophagic flux, attenuating weight gain, enhancing DC, T-cell and NK cell numbers, and promoting long-term tumor control in a murine hepatic metastases model. Autophagy (programmed cell survival) is a metabolic process associated with promotion of late cancer growth. In tumor cell lines, CQ treatment limits ATP production through inhibition of oxidative phosphorylation and promotion of apoptosis. CQ increases autophagic vacuoles and LC3-II levels in tumor cells, associated with increased annexin V⁺/PI^- cells, cleaved-PARP, cleaved-CASP3, and cytochrome c release from mitochondria. These observations, limiting toxicity and prolonging antitumor effects, with a combination of IL2 and autophagy inhibition in murine models are now being tested by the Cytokine Working Group in patients with advanced renal cell carcinoma.     

Induction of autophagy during extracellular matrix detachment promotes cell survival

Autophagy has been proposed to promote cell death during lumen formation in three-dimensional mammary epithelial acini because numerous autophagic vacuoles are observed in the dying central cells during morphogenesis. Because these central cells die due to extracellular matrix (ECM) deprivation (anoikis), we have directly interrogated how matrix detachment regulates autophagy. Detachment induces autophagy in both nontumorigenic epithelial lines and in primary epithelial cells. RNA interference-mediated depletion of autophagy regulators (ATGs) inhibits detachment-induced autophagy, enhances apoptosis, and reduces clonogenic recovery after anoikis. Remarkably, matrix-detached cells still exhibit autophagy when apoptosis is blocked by Bcl-2 overexpression, and ATG depletion reduces the clonogenic survival of Bcl-2-expressing cells after detachment. Finally, stable reduction of ATG5 or ATG7 in MCF-10A acini enhances luminal apoptosis during morphogenesis and fails to elicit long-term luminal filling, even when combined with apoptotic inhibition mediated by Bcl-2 overexpression. Thus, autophagy promotes epithelial cell survival during anoikis, including detached cells harboring antiapoptotic lesions.     

细胞自噬与肿瘤耐药

细胞自噬是一种细胞自我降解的过程,在适应代谢应激、保持基因组完整性及维持内环境稳定方面发挥重要作用. 在肿瘤治疗中,凋亡耐受是产生肿瘤耐药的重要机制. 细胞自噬可防止抗肿瘤药诱导的凋亡,促进肿瘤耐药. 然而,自噬性细胞死亡可能是凋亡耐受肿瘤细胞的一种死亡方式. 因此,细胞自噬对肿瘤细胞的耐药性有双重影响. 本文综述了细胞自噬的分子机制、细胞自噬与凋亡的关系、细胞自噬与肿瘤耐药以及治疗的主要研究进展.     

Autophagy as a trigger for cell death: autophagic degradation of inhibitor of apoptosis dBruce controls DNA fragmentation during late oogenesis in Drosophila

Autophagy has been reported to contribute to cell death, but the underlying mechanisms remain largely unknown and controversial. We have: been studying oogenesis in Drosophila melanogaster as a model system to understand the interplay between autophagy and cell death. Using a novel autophagy reporter we found that autophagy occurs during developmental cell death of nurse cells in late oogenesis. Genetic inhibition: of autophagy-related genes atg1, atg13 and vps34 results in late-stage egg chambers containing persisting nurse cell nuclei without fragmented DNA and attenuation of caspase-3 cleavage. We found that Drosophila inhibitor of apoptosis dBruce is degraded by autophagy and this degradation promotes DNA fragmentation and subsequent nurse cell death. These studies demonstrate that autophagic degradation of an inhibitor: of apoptosis is a novel mechanism of triggering cell death.     

An emerging role for nanomaterials in increasing immunogenicity of cancer cell death

In the last decade, it has become clear that anti-cancer therapy is more successful when it can also induce an immunogenic form of cancer cell death (ICD). ICD is an umbrella term covering several cell death modalities, including apoptosis and necroptosis. In general, ICD is characterized by the emission of damage-associated molecular patterns (DAMPs) and/or cytokines/chemokines, leading to the induction of strong anti-tumor immune responses. In experimental cancer therapy, new observations indicate that the immunogenicity of dying cancer cells can be improved by the use of biomaterials. In this review, after a brief overview of the basic principles of the concept of ICD and discussion of the potential use of DAMPs as biomarkers of therapy efficacy, we discuss an emerging role of nanomaterials as a promising strategy to modulate the immunogenicity of cancer cell death. We address how nanocarriers can be used to increase the immunogenicity of ICD and then turn our attention to their dual action. Nanocarriers can be used to increase the immunogenicity of dying cancer cells and to reduce the side effects of chemotherapy. Future studies will show whether biomaterials are truly an optimal strategy to modulate the immunogenicity of dying cancer cells and will provide the insights needed for the development of novel treatment strategies for cancer.     

Autophagy in the pathogenesis of myelodysplastic syndrome and acute myeloid leukemia

Autophagy is a conserved cellular pathway responsible for the sequestration of spent organelles and protein aggregates from the cytoplasm and their delivery into lysosomes for degradation. Autophagy plays an important role in adaptation to starvation, in cell survival, immunity, development and cancer. Recent evidence in mice suggests that autophagic defects in hematopoietic stem cells (HSCs) may be implicated in leukemia. Indeed, mice lacking Atg7 in HSCs develop an atypical myeloproliferation resembling human myelodysplastic syndrome (MDS) progressing to acute myeloid leukemia (AML). Studies suggest that accumulation of damaged mitochondria and reactive oxygen species result in cell death of the majority of progenitor cells and, possibly, concomitant transformation of some surviving ones. Interestingly, bone marrow cells from MDS patients are characterized by mitochondrial abnormalities and increased cell death. A role for autophagy in the transformation to cancer has been proposed in other cancer types. This review focuses on autophagy in human MDS development and progression to AML within the context of the role of mitochondria, apoptosis and reactive oxygen species (ROS) in its pathogenesis.     

Autophagy in the pathogenesis of myelodysplastic syndrome and acute myeloid leukemia

Autophagy is a conserved cellular pathway responsible for the sequestration of spent organelles and protein aggregates from the cytoplasm and their delivery into lysosomes for degradation. Autophagy plays an important role in adaptation to starvation, in cell survival, immunity, development and cancer. Recent evidence in mice suggests that autophagic defects in hematopoietic stem cells (HSCs) may be implicated in leukemia. Indeed, mice lacking Atg7 in HSCs develop an atypical myeloproliferation resembling human myelodysplastic syndrome (MDS) progressing to acute myeloid leukemia (AML). Our studies suggest that accumulation of damaged mitochondria and reactive oxygen species result in cell death of the majority of progenitor cells and, possibly, concomitant transformation of some surviving ones. Interestingly, bone marrow cells from MDS patients are characterized by mitochondrial abnormalities and increased cell death. A role for autophagy in the transformation to cancer has been proposed in other cancer types. This review focuses on autophagy in human MDS development and progression to AML within the context of the role of mitochondria, apoptosis and reactive oxygen species (ROS) in its pathogenesis.Key words: autophagy, mitophagy, Atg7, hematopoiesis, HSCs, myelodysplastic syndrome, acute myeloid leukemia     

Autophagic degradation of GZMB/granzyme B

The crucial issue for defining successful natural killer (NK)-based anticancer therapy is the ability of tumor cells to activate resistance mechanisms leading to escape from NK-mediated killing. It is now well established that such mechanisms are likely evolved under hypoxia in the tumor microenvironment. Here, we show that hypoxia-induced autophagy impairs breast cancer cell susceptibility to NK-mediated lysis and that this impairment is reverted by targeting autophagy. We provide evidence that activation of autophagy in hypoxic cells is involved in selective degradation of the pro-apoptotic NK-derived serine protease GZMB/granzyme B, thereby blocking NK-mediated target cell apoptosis. Our in vivo data validate the concept that targeting autophagy in cancer cells promotes tumor regression by facilitating their elimination by NK cells. This study provides a cutting-edge advance in our understanding of how hypoxia-induced autophagy impairs NK-mediated lysis and might pave the way for formulating more effective NK-based antitumor therapy by combining autophagy inhibitors.     

Commentary: Autophagic degradation of GZMB/granzyme B: A new mechanism of hypoxic tumor cell escape from natural killer cell-mediated lysis

The crucial issue for defining successful natural killer (NK)-based anticancer therapy is the ability of tumor cells to activate resistance mechanisms leading to escape from NK-mediated killing. It is now well established that such mechanisms are likely evolved under hypoxia in the tumor microenvironment. Here, we show that hypoxia-induced autophagy impairs breast cancer cell susceptibility to NK-mediated lysis and that this impairment is reverted by targeting autophagy. We provide evidence that activation of autophagy in hypoxic cells is involved in selective degradation of the pro-apoptotic NK-derived serine protease GZMB/granzyme B, thereby blocking NK-mediated target cell apoptosis. Our in vivo data validate the concept that targeting autophagy in cancer cells promotes tumor regression by facilitating their elimination by NK cells. This study provides a cutting-edge advance in our understanding of how hypoxia-induced autophagy impairs NK-mediated lysis and might pave the way for formulating more effective NK-based antitumor therapy by combining autophagy inhibitors.     

Autophagy induction impairs Wnt/β-catenin signalling through β-catenin relocalisation in glioblastoma cells

Autophagy is an evolutionary conserved process mediating lysosomal degradation of cytoplasmic material. Its involvement in cancer progression is highly controversial, due to its dual role in both limiting tumoural transformation and in protecting established tumoral cells from unfavorable conditions. Little is known about the cross-talk between autophagy and intracellular signalling pathways, as well as about autophagy impact on signalling molecules turnover.An aberrantly activated Wnt/β-catenin signalling is responsible for tumour proliferation, invasion, and stemness maintenance. Here we show that autophagy negatively regulates Wnt/β-catenin signalling in glioblastoma multiforme (GBM) cells, through Dishevelled degradation. We also provide the first evidence that autophagy promotes β-catenin relocalisation within the cell, by inducing a decrease of the nuclear protein fraction. In particular, upon autophagy induction, β-catenin appears mainly localized in sub-membrane areas where it associates with N-cadherin to form epithelial-like cell-cell adhesion structures.Our data indicate, for the first time, that autophagy induction results in Wnt signalling attenuation and in β-catenin relocalisation within the GBM cell. These findings further support the idea that autophagy modulation could represent a potential therapeutical strategy to contrast GBM progression.     

Three dimensions of autophagy in regulating tumor growth: cell survival/death,cell proliferation,and tumor dormancy

Autophagy is an intracellular lysosomal degradation process involved in multiple facets of cancer biology. Various dimensions of autophagy are associated with tumor growth and cancer progression, and here we focus on the dimensions involved in regulation of cell survival/cell death, cell proliferation and tumor dormancy. The first dimension of autophagy supports cell survival under stress within tumors and under certain contexts drives cell death, impacting tumor growth. The second dimension of autophagy promotes proliferation through directly regulating cell cycle or indirectly maintaining metabolism, increasing tumor growth. The third dimension of autophagy facilitates tumor cell dormancy, contributing to cancer treatment resistance and cancer recurrence. The intricate relationship between these three dimensions of autophagy influences the extent of tumor growth and cancer progression. In this review, we summarize the roles of the three dimensions of autophagy in tumor growth and cancer progression, and discuss unanswered questions in these fields.     

Mechanisms of autophagy and apoptosis: Recent developments in breast cancer cells

Autophagy,the pathway whereby cell components are degraded by lysosomes,is involved in the cell response to environmental stresses,such as nutrient deprivation,hypoxia or exposition to chemotherapeutic agents.Under these conditions,which are reminiscent of certain phases of tumor development,autophagy either promotes cell survival or induces cell death. This strengthens the possibility that autophagy could be an important target in cancer therapy,as has been proposed.Here,we describe the regulation of survival and death by autophagy and apoptosis,especially in cultured breast cancer cells.In particular,we discuss whether autophagy represents an apoptosis-independent process and/or if they share common pathways. We believe that understanding in detail the molecular mechanisms that underlie the relationships between autophagy and apoptosis in breast cancer cells could improve the available treatments for this disease.