NRF2-driven redox metabolism takes center stage in cancer metabolism from an outside-in perspective

Cancer development is a process of somatic clonal evolution. Darwinian principles of evolution emphasize the interaction between heritable individual variability and selective pressure from the environment. However, the current prevailing concept of cancer evolution mostly focuses on the alterations of genes, signaling, and metabolism inside cells, which underestimates the impact of environmental pressure in selecting the adapted cells. Recently, unsuccessful outcomes and many concerns raised in targeting those alterations inside cells have cast doubt on the current “cell-centric” paradigm of cancer formation, which necessitates a paradigm shift to an outside-in direction that considers environmental changes as a driver in determining the characteristics of selected cells. In the tumor microenvironment, reactive oxygen species (ROS) are one of the most abundant chemical constituents generated by inflammatory and hypoxic conditions. Because of their cytotoxicity when present at high levels, ROS should be the pressure that selects cells with a high capacity for ROS metabolism and antioxidant defense, both of which are referred to as redox metabolism. Cancer genome analyses have found that nuclear factor E2-related factor 2 (NRF2), which plays an indispensable role in redox metabolism, is frequently activated in many types of cancer, particularly lung cancer. This suggests that an ROS-rich microenvironment drives the selection, survival, and growth of cells with high NRF2 activity. Thus, NRF2-driven redox metabolism should be the most crucial part of cancer metabolism, proposing NRF2 inhibitor as an attractive therapeutic target for cancer.

     

Reactive oxygen species as double-edged swords in cellular processes: low-dose cell signaling versus high-dose toxicity

ROS are diverse and abundant in biological systems. While excessive ROS production clearly damages DNA, low levels of ROS affect cell signaling particularly at the level of redox modulation. Moreover, the specific contributions of ROS to apoptosis and mitogenesis in maintenance of cell number homeostasis remains to be elucidated. ROS dose is a critical parameter in determining the ultimate cellular response; however the shape of the dose response curve is unpredictable. When cells are stimulated with ROS, cell-signaling cascades are activated. It appears that the cellular redox potential is an important determinant of cell function and interruption of redox balance may adversely affect cell function. As a result, compounds such as antioxidants may intercept critical ROS signaling molecules and both protect cells and foster pathogenesis. As a result, further study is needed to unravel the role of ROS in redox regulation and the potential outcome of antioxidant administration on cellular responses.     

Redox-active quinones and ascorbate: an innovative cancer therapy that exploits the vulnerability of cancer cells to oxidative stress

Cancer cells are particularly vulnerable to treatments impairing redox homeostasis. Reactive oxygen species (ROS) can indeed play an important role in the initiation and progression of cancer, and advanced stage tumors frequently exhibit high basal levels of ROS that stimulate cell proliferation and promote genetic instability. In addition, an inverse correlation between histological grade and antioxidant enzyme activities is frequently observed in human tumors, further supporting the existence of a redox dysregulation in cancer cells. This biochemical property can be exploited by using redox-modulating compounds, which represent an interesting approach to induce cancer cell death. Thus, we have developed a new strategy based on the use of pharmacologic concentrations of ascorbate and redox-active quinones. Ascorbate-driven quinone redox cycling leads to ROS formation and provoke an oxidative stress that preferentially kill cancer cells and spare healthy tissues. Cancer cell death occurs through necrosis and the underlying mechanism implies an energetic impairment (ATP depletion) that is likely due to glycolysis inhibition. Additional mechanisms that participate to cell death include calcium equilibrium impairment and oxidative cleavage of protein chaperone Hsp90. Given the low systemic toxicity of ascorbate and the impairment of crucial survival pathways when associated with redox-active quinones, these combinations could represent an original approach that could be combined to standard cancer therapy.     

Experimental therapeutics: targeting the redox Achilles heel of cancer

Reactive oxygen species (ROS) have recently emerged as promising targets for anticancer drug discovery. Constitutively elevated levels of cellular oxidative stress and dependence on mitogenic and anti-apoptotic ROS signaling represent a specific vulnerability of malignant cells that can be selectively targeted by novel pro- and antioxidant redox chemotherapeutics. This review discusses small-molecule anticancer redox drugs currently in various phases of preclinical and clinical development that are characterized by their unique mechanism of action, including small-molecule superoxide dismutase and catalase mimetics, bioreductively activated pro-oxidant redox catalysts, metal-based pro-oxidants, hypoxia-selective free radical precursors, and specific antagonists of the cancer cell antioxidant glutathione or thioredoxin redox systems. Based on ongoing redox biomarker discovery and validation, future redox phenotyping and genotyping may guide the selection of novel redox chemotherapeutics that efficiently target the redox Achilles heel of the individual tumor.     

Cellular redox pathways as a therapeutic target in the treatment of cancer

The vulnerability of some cancer cells to oxidative signals is a therapeutic target for the rational design of new anticancer agents. In addition to their well characterized effects on cell division, many cytotoxic anticancer agents can induce oxidative stress by modulating levels of reactive oxygen species (ROS) such as the superoxide anion radical, hydrogen peroxide and hydroxyl radicals. Tumour cells are particularly sensitive to oxidative stress as they typically have persistently higher levels of ROS than normal cells due to the dysregulation of redox balance that develops in cancer cells in response to increased intracellular production of ROS or depletion of antioxidant proteins. In addition, excess ROS levels potentially contribute to oncogenesis by the mediation of oxidative DNA damage. There are several anticancer agents in development that target cellular redox regulation. The overall cellular redox state is regulated by three systems that modulate cellular redox status by counteracting free radicals and ROS, or by reversing the formation of disulfides; two of these are dependent on glutathione and the third on thioredoxin. Drugs targeting S-glutathionylation have direct anticancer effects via cell signalling pathways and inhibition of DNA repair, and have an impact on a wide range of signalling pathways. Of these agents, NOV-002 and canfosfamide have been assessed in phase III trials, while a number of others are undergoing evaluation in early phase clinical trials. Alternatively, agents including PX-12, dimesna and motexafin gadolinium are being developed to target thioredoxin, which is overexpressed in many human tumours, and this overexpression is associated with aggressive tumour growth and poorer clinical outcomes. Finally, arsenic derivatives have demonstrated antitumour activity including antiproliferative and apoptogenic effects on cancer cells by pro-oxidant mechanisms, and the induction of high levels of oxidative stress and apoptosis by an as yet undefined mechanism. In this article we review anticancer drugs currently in development that target cellular redox activity to treat cancer.     

Pro-oxidant natural products as anticancer agents

Cancer cells produce high levels of reactive oxygen species (ROS) that lead to a state of increased basal oxidative stress. Since this state of oxidative stress makes cancer cells vulnerable to agents that further augment ROS levels, the use of pro-oxidant agents is emerging as an exciting strategy to selectively target tumor cells. Natural products have provided a significant contribution to the development of several drugs currently used in cancer chemotherapy. Although many natural products are known to affect the redox state of the cell, most studies on these compounds have focused on their antioxidant activity instead of on their pro-oxidant properties. This article provides an overview of natural products with pro-oxidant and anticancer activities, with special focus on plant secondary metabolites, and discusses their possible use as cancer chemotherapeutic agents.     

Targeting the Fas/FasL signaling pathway in cancer therapy

INTRODUCTION: The Fas/FasL system plays a significant role in tumorigenesis. Research has shown that its impairment in cancer cells may lead to apoptosis resistance and contribute to tumor progression. Thus, the development of effective therapies targeting the Fas/FasL system may play an important role in the fight against cancer. AREAS COVERED: In this review the recent literature on targeting the Fas/FasL system for therapeutic exploitation at different levels is reviewed. Promising pre-clinical approaches and various exceptions are highlighted. The potential of combined therapies is also explored, whereby tumor sensitivity to Fas-mediated apoptosis is restored, before an effective targeted therapy is employed. EXPERT OPINION: The success of the Fas/FasL system targeting for therapeutics will require a better understanding of the alterations conferring resistance, in order to use the most appropriate sensitizing chemotherapeutic or radiotherapeutic agents in combination with effective targeted therapies.     

PARP and other prospective targets for poisoning cancer cell metabolism

Increasing evidence indicates that cancer cells rewire their metabolism during tumorigenesis. The high intracellular levels of lactate and reactive oxygen species (ROS) generated during enhanced aerobic glycolysis and mitochondrial oxidative phosphorylation respectively led to oxidative stress. The detoxification of these accumulating metabolites and the equilibrium between reduced and oxidized nicotine adenine dinucleotide (NADH and NAD⁺) are two prominent mechanisms regulating redox status and hence energy homeostasis in tumors. Targeting both processes may thus be selectively cytotoxic for cancer cells. In this context, the impact of poly(ADP-ribose) polymerase (PARP) inhibitors, a class of anticancer agents employed for the treatment of DNA repair deficient tumors, on energy homeostasis and mitochondrial respiration regulation has potential clinical implications. Here we provide an overview of the metabolic reprogramming occurring in cancer cells and discuss the translational perspectives of targeting tumor metabolism and redox balance for antineoplastic therapy.     

Effect of selenium-saturated bovine lactoferrin (Se-bLF) on antioxidant enzyme activities in human gut epithelial cells under oxidative stress

Cancer and many chronic inflammatory diseases are associated with increased amounts of reactive oxygen species (ROS). The potential cellular and tissue damage created by ROS has significant impact on many disease and cancer states and natural therapeutics are becoming essential in regulating altered redox states. We have shown recently that iron content is a critical determinant in the antitumour activity of bovine milk lactoferrin (bLF). We found that 100% iron-saturated bLF (Fe-bLF) acts as a potent natural adjuvant and fortifying agent for augmenting cancer chemotherapy and thus has a broad utility in the treatment of cancer. Furthermore, we also studied the effects of iron saturated bLF's ability as an antioxidant in the human epithelial colon cancer cell line HT29, giving insights into the potential of bLF in its different states. Thus, metal saturated bLF could be implemented as anti-cancer neutraceutical. In this regard, we have recently been able to prepare a selenium (Se) saturated form of bLF, being up to 98% saturated. Therefore, the objectives of this study were to determine how oxidative stress induced by hydrogen peroxide (H2O2) alters antioxidant enzyme activity within HT29 epithelial colon cancer cells, and observe changes in this activity by treatments with different antioxidants ascorbic acid (AA), Apo (iron free)-bLF and selenium (Se)-bLF. The states of all antioxidant enzymes (glutathione peroxidase (GPx), glutathione reductase (GR), glutathione- s-transferase (GsT), catalase and superoxide dismutase (SOD)) demonstrated high levels within untreated HT29 cells compared to the majority of other treatments being used, even prior to H2O2 exposure. All enzymes showed significant alterations in activity when cells were treated with antioxidants AA, Apo-bLF or Se-bLF, with and/or without H2O2 exposure. Obvious indications that the Se content of the bLF potentially interacted with the glutathione (GSH)/GPx/GR/GsT associated redox system could be observed immediately, showing capability of Se-bLF being highly beneficial in helping to maintain a balance between the oxidant/antioxidant systems within cells and tissues, especially in selenium deficient systems. In conclusion, the antioxidative defence activity of Se-bLf, investigated in this study for the first time, shows dynamic adaptations that may allow for essential protection from the imbalanced oxidative conditions. Because of its lack of toxicity and the availability of both selenium and bLF in whole milk, Se-bLF offers a promise for a prospective natural dietary supplement, in addition to being an immune system enhancement, or a potential chemopreventive agent for cancers.     

An epigrammatic (abridged) recounting of the myriad tales of astonishing deeds and dire consequences pertaining to nitric oxide and reactive oxygen species in mitochondria with an ancillary missive concerning the origins of apoptosis

Mitochondria play a central role in the life and death of cells. These organelles serve as the major energy-producing power-house, whereby the generation of ATP is associated with the utilization of molecular oxygen. A significant fraction (2-3%) of molecular oxygen consumed by mitochondria may be reduced in a one-electron fashion to yield a series of reactive oxygen species (ROS) such as superoxide anion radical, hydrogen peroxide, and hydroxyl radical. ROS are capable of damaging components of the electron transport apparatus and can, in turn, disrupt mitochondrial functioning, limiting cellular ATP levels and ultimately resulting in cell death. ROS-induced disruption of electron transport can perpetuate production of deleterious ROS and propagate mitochondrial damage. Consequently, mitochondria are highly enriched with water-soluble and lipid-soluble antioxidants (glutathione, ascorbate, Vitamin E, and coenzyme Q) and antioxidant enzymes, such as superoxide dismutase, glutathione peroxidase, catalase, thioredoxins, and peroxiredoxin. Another important antioxidant acting as a very effective scavenger of reactive lipid (peroxyl, alkoxyl) radicals is nitric oxide (NO) generated by mitochondrial nitric oxide synthase. However, NO can also be very disruptive to mitochondria function, a process facilitated by its high reactivity with superoxide. This interaction results in the formation of peroxynitrite, an oxidant capable of causing oxidative/nitrosative stress, further aggravating mitochondrial dysfunction, causing ATP depletion and damage to cells. Thus, in the most general sense, the effects of NO in mitochondria may be either protective or deleterious depending on specific conditions of local redox environment (redox potential, ratio of oxidized to reduced glutathione, transition metals, and the presence of other oxygen- and nitrogen-centered radicals).     

Sodium arsenite induces cyclooxygenase-2 expression in human uroepithelial cells through MAPK pathway activation and reactive oxygen species induction

Arsenic can induce reactive oxygen species (ROS) leading to oxidative stress and carcinogenesis. Bladder is one of the major target organs of arsenic, and cyclooxygenase-2 (COX-2) may play an important role in arsenic-induced bladder cancer. However, the mechanism by which arsenic induces COX-2 in bladder cells remains unclear. This study aimed at investigating arsenic-mediated intracellular redox status and signaling cascades leading to COX-2 induction in human uroepithelial cells (SV-HUC-1). SV-HUC-1 cells were exposed to sodium arsenite and COX-2 expression, mitogen-activated protein kinase (MAPK) phosphorylation, glutathione (GSH) levels, ROS induction and Nrf2 expression were quantified. Our results demonstrate that arsenite (1–10 μM) elevates COX-2 expression, GSH levels, ROS and Nrf2 expression. Arsenite treatment for 24 h stimulates phosphorylation of ERK and p38, but not JNK in SV-HUC-1 cells. Induction of Cox-2 mRNA levels by arsenite was attenuated by inhibitors of ERK, p38 and JNK. Arsenite-induced ROS generation and COX-2 expression were significantly attenuated by treatment with melatonin (a ROS scavenger), but enhanced by DL-buthionine-(S, R)-sulfoximine (BSO, an inhibitor of gamma-glutamylcysteine synthetase (γ-GCS) resulting in lower GSH and increased ROS levels). These data indicate that arsenite promotes an induction of ROS, which results in an induction of COX-2 expression through activation of the MAPK pathway.     

Tumor intracellular redox status and drug resistance--serendipity or a causal relationship?

Reducing tumor load by therapeutic induction of cell death in the transformed phenotype is the desirable goal of most chemotherapeutic regimens. Despite the tremendous strides made in our understanding of mechanisms that endow tumor cells with the ability to evade execution signals, development of chemo-resistance is still a major obstacle in the successful management of the disease. A host of factors have been implicated in the acquisition of the resistant phenotype, such as activation of drug efflux pumps, overexpression of proteins that inhibit cell death, absence of critical members of the death circuitry, and selective loss of cell cycle checkpoints. Consequently, it is now well established that the process of carcinogenesis is not only a result of an increase in cells' proliferative capacity, but a product of increased proliferation and defective or diminished cell death signaling. To that end, one of the critical determinants of cellular response to exogenous stimuli is the cellular redox status. Intracellular generation of reactive oxygen species (ROS) is tightly regulated by the intrinsic anti-oxidant defense systems. Despite the conventional dogma that ROS are harmful to the cell, experimental evidence over the last decade or so bear witness to the fact that ROS also play an important role as signaling molecules in diverse physiological processes. Indeed, low levels of intracellular ROS have been linked to cellular proliferation and cell cycle progression, which provides an explanation for the pro-oxidant state invariably associated with the transformed phenotype. Coupled to that are recent observations implicating pro-oxidant intracellular milieu in tumor cells' resistance to cell death signals delivered through the cell surface receptor or upon exposure to chemotherapeutic drugs. These studies provide convincing evidence to support a direct or indirect role for intracellular superoxide anion in creating an intracellular milieu non-permissive for cell death execution. Thus a novel approach to enhancing tumor cell sensitivity to chemotherapy-induced cell death would be to favourably tailor the cytosolic milieu to allow efficient apoptotic execution. Here we present a brief discussion on the role of ROS in cell growth and differentiation, and more specifically address the issue of chemo-resistance from the standpoint of cellular redox status.     

Mitigation of Cu(II)-induced damage in human blood cells by carnosine: An in vitro study

Copper (Cu) is an essential micronutrient but human exposure to high level of this metal results in adverse health effects. Oxidative stress is assumed to play a major role in the mechanism of Cu-induced toxicity. The protective role of carnosine, an antioxidant and antiglycating agent, was examined against Cu-induced toxicity in isolated human blood cells. Red blood cells (RBC) were treated with 0.5 mM copper chloride (CuCl₂), a Cu(II) compound, either alone or after treatment with carnosine. Incubation of RBC with CuCl₂ increased protein oxidation, lipid peroxidation, methemoglobin formation and lowered glutathione content. The antioxidant defense system was impaired and production of reactive oxygen (ROS) and reactive nitrogen species (RNS) was enhanced. Pre-incubation of RBC with carnosine protected the cells against CuCl₂-induced oxidative damage. It restored the activities of several antioxidant, membrane-bound and metabolic enzymes, decreased the generation of ROS and RNS, enhanced the antioxidant power of cells and prevented inactivation of plasma membrane redox system. Carnosine also protected human lymphocytes from CuCl₂-induced DNA damage. The protective effects of carnosine were concentration-dependent while carnosine itself did not exhibit any adverse effect. Carnosine can, therefore, be used as a possible chemoprotectant against the harmful effects of this extremely redox active metal.     

Divergent Effects of Sulforaphane on Basal and Glucose-Stimulated Insulin Secretion in β-Cells: Role of Reactive Oxygen Species and Induction of Endogenous Antioxidants

Purpose

Oxidative stress is implicated in pancreatic β-cell dysfunction, yet clinical outcomes of antioxidant therapies on diabetes are inconclusive. Since reactive oxygen species (ROS) can function as signaling intermediates for glucose-stimulated insulin secretion (GSIS), we hypothesize that exogenously boosting cellular antioxidant capacity dampens signaling ROS and GSIS.

Methods

To test the hypothesis, we formulated a mathematical model of redox homeostatic control circuit comprising known feedback and feedforward loops and validated model predictions with plant-derived antioxidant sulforaphane (SFN).

Results

SFN acutely (30-min treatment) stimulated basal insulin secretion in INS-1(832/13) cells and cultured mouse islets, which could be attributed to SFN-elicited ROS as N-acetylcysteine or glutathione ethyl ester suppressed SFN-stimulated insulin secretion. The mathematical model predicted an adapted redox state characteristic of strong induction of endogenous antioxidants but marginally increased ROS under prolonged SFN exposure, a state that attenuates rather than facilitates glucose-stimulated ROS and GSIS. We validated the prediction by demonstrating that although 24-h treatment of INS-1(832/13) cells with low, non-cytotoxic concentrations of SFN (2–10 μM) protected the cells from cytotoxicity by oxidative insult, it markedly suppressed insulin secretion stimulated by 20 mM glucose.

Conclusions

Our study indicates that adaptive induction of endogenous antioxidants by exogenous antioxidants, albeit cytoprotective, inhibits GSIS in β-cells.