Iron Chelator Induces Apoptosis in Osteosarcoma Cells by Disrupting Intracellular Iron Homeostasis and Activating the MAPK Pathway

Osteosarcoma is a common malignant bone tumor in clinical orthopedics. Iron chelators have inhibitory effects on many cancers, but their effects and mechanisms in osteosarcoma are still uncertain. Our in vitro results show that deferoxamine (DFO) and deferasirox (DFX), two iron chelators, significantly inhibited the proliferation of osteosarcoma cells (MG-63, MNNG/HOS and K7M2). The viability of osteosarcoma cells was decreased by DFO and DFX in a concentration-dependent manner. DFO and DFX generated reactive oxygen species (ROS), altered iron metabolism and triggered apoptosis in osteosarcoma cells. Iron chelator-induced apoptosis was due to the activation of the MAPK signaling pathway, with increased phosphorylation levels of JNK, p38 and ERK, and ROS generation; in this process, the expression of C-caspase-3 and C-PARP increased. In an orthotopic osteosarcoma transplantation model, iron chelators (20 mg/kg every day, Ip, for 14 days) significantly inhibited the growth of the tumor. Immunohistochemical analysis showed that iron metabolism was altered, apoptosis was promoted, and malignant proliferation was reduced with iron chelators in the tumor tissues. In conclusion, we observed that iron chelators induced apoptosis in osteosarcoma by activating the ROS-related MAPK signaling pathway. Because iron is crucial for cell proliferation, iron chelators may provide a novel therapeutic strategy for osteosarcoma.     

Recent Advances in Iron Chelation and Gallium-Based Therapies for Antibiotic Resistant Bacterial Infections

Iron is essential for multiple bacterial processes and is thus required for host colonization and infection. The antimicrobial activity of multiple iron chelators and gallium-based therapies against different bacterial species has been characterized in preclinical studies. In this review, we provide a synthesis of studies characterizing the antimicrobial activity of the major classes of iron chelators (hydroxamates, aminocarboxylates and hydroxypyridinones) and gallium compounds. Special emphasis is placed on recent in-vitro and in-vivo studies with the novel iron chelator DIBI. Limitations associated with iron chelation and gallium-based therapies are presented, with emphasis on limitations of preclinical models, lack of understanding regarding mechanisms of action, and potential host toxicity. Collectively, these studies demonstrate potential for iron chelators and gallium to be used as antimicrobial agents, particularly in combination with existing antibiotics. Additional studies are needed in order to characterize the activity of these compounds under physiologic conditions and address potential limitations associated with their clinical use as antimicrobial agents.     

The Iron Chelator,Dp44mT,Effectively Inhibits Human Oral Squamous Cell Carcinoma Cell Growth in Vitro and in Vivo

Oral squamous cell carcinoma (OSCC) is a common malignancy with a growing worldwide incidence and prevalence. The N-myc downstream regulated gene (NDRG) family of NDRG1, 2, 3, and mammary serine protease inhibitor (Maspin) gene are well-known modulators in the neoplasia process. Current research has considered iron chelators as new anti-cancer agents; however, the anticancer activities of iron chelators and their target genes in OSCC have not been well investigated. We showed that iron chelators (Dp44mT, desferrioxamine (DFO), and deferasirox) all significantly inhibit SAS cell growth. Flow cytometry further indicated that Dp44mT inhibition of SAS cells growth was partly due to induction of G1 cell cycle arrest. Iron chelators enhanced expressions of NDRG1 and NDRG3 while repressing cyclin D1 expression in OSCC cells. The in vivo antitumor effect on OSCC and safety of Dp44mT were further confirmed through a xenograft animal model. The Dp44mT treatment also increased Maspin protein levels in SAS and OECM-1 cells. NDRG3 knockdown enhanced the growth of OECM-1 cells in vitro and in vivo. Our results indicated that NDRG3 is a tumor suppressor gene in OSCC cells, and Dp44mT could be a promising therapeutic agent for OSCC treatment.     

Emerging Roles of the Iron Chelators in Inflammation

Iron is a crucial element for mammalian cells, considering its intervention in several physiologic processes. Its homeostasis is finely regulated, and its alteration could be responsible for the onset of several disorders. Iron is closely related to inflammation; indeed, during inflammation high levels of interleukin-6 cause an increased production of hepcidin which induces a degradation of ferroportin. Ferroportin degradation leads to decreased iron efflux that culminates in elevated intracellular iron concentration and consequently iron toxicity in cells and tissues. Therefore, iron chelation could be considered a novel and useful therapeutic strategy in order to counteract the inflammation in several autoimmune and inflammatory diseases. Several iron chelators are already known to have anti-inflammatory effects, among them deferiprone, deferoxamine, deferasirox, and Dp44mT are noteworthy. Recently, eltrombopag has been reported to have an important role in reducing inflammation, acting both directly by chelating iron, and indirectly by modulating iron efflux. This review offers an overview of the possible novel biological effects of the iron chelators in inflammation, suggesting them as novel anti-inflammatory molecules.     

Is Chelation Therapy a Potential Treatment for Parkinson’s Disease?

Iron loading in some brain regions occurs in Parkinson’s Disease (PD), and it has been considered that its removal by iron chelators could be an appropriate therapeutic approach. Since neuroinflammation with microgliosis is also a common feature of PD, it is possible that iron is sequestered within cells as a result of the “anaemia of chronic disease” and remains unavailable to the chelator. In this review, the extent of neuroinflammation in PD is discussed together with the role played by glia cells, specifically microglia and astrocytes, in controlling iron metabolism during inflammation, together with the results of MRI studies. The current use of chelators in clinical medicine is presented together with a discussion of two clinical trials of PD patients where an iron chelator was administered and showed encouraging results. It is proposed that the use of anti-inflammatory drugs combined with an iron chelator might be a better approach to increase chelator efficacy.     

Management of Iron Overload in Beta-Thalassemia Patients: Clinical Practice Update Based on Case Series

Thalassemia syndromes are characterized by the inability to produce normal hemoglobin. Ineffective erythropoiesis and red cell transfusions are sources of excess iron that the human organism is unable to remove. Iron that is not saturated by transferrin is a toxic agent that, in transfusion-dependent patients, leads to death from iron-induced cardiomyopathy in the second decade of life. The availability of effective iron chelators, advances in the understanding of the mechanism of iron toxicity and overloading, and the availability of noninvasive methods to monitor iron loading and unloading in the liver, heart, and pancreas have all significantly increased the survival of patients with thalassemia. Prolonged exposure to iron toxicity is involved in the development of endocrinopathy, osteoporosis, cirrhosis, renal failure, and malignant transformation. Now that survival has been dramatically improved, the challenge of iron chelation therapy is to prevent complications. The time has come to consider that the primary goal of chelation therapy is to avoid 24-h exposure to toxic iron and maintain body iron levels within the normal range, avoiding possible chelation-related damage. It is very important to minimize irreversible organ damage to prevent malignant transformation before complications set in and make patients ineligible for current and future curative therapies. In this clinical case-based review, we highlight particular aspects of the management of iron overload in patients with beta-thalassemia syndromes, focusing on our own experience in treating such patients. We review the pathophysiology of iron overload and the different ways to assess, quantify, and monitor it. We also discuss chelation strategies that can be used with currently available chelators, balancing the need to keep non-transferrin-bound iron levels to a minimum (zero) 24 h a day, 7 days a week and the risk of over-chelation.     

Binding of Citrate-Fe3+ to Plastic Culture Dishes,an Artefact Useful as a Simple Technique to Screen for New Iron Chelators

NaCT mediates citrate uptake in the liver cell line HepG2. When these cells were exposed to iron (Fe³⁺), citrate uptake/binding as monitored by the association of [¹⁴C]-citrate with cells increased. However, there was no change in NaCT expression and function, indicating that NaCT was not responsible for this Fe³⁺-induced citrate uptake/binding. Interestingly however, the process exhibited substrate selectivity and saturability as if the process was mediated by a transporter. Notwithstanding these features, subsequent studies demonstrated that the iron-induced citrate uptake/binding did not involve citrate entry into cells; instead, the increase was due to the formation of citrate-Fe³⁺ chelate that adsorbed to the cell surface. Surprisingly, the same phenomenon was observed in culture wells without HepG2 cells, indicating the adsorption of the citrate-Fe³⁺ chelate to the plastic surface of culture wells. We used this interesting phenomenon as a simple screening technique for new iron chelators with the logic that if another iron chelator is present in the assay system, it would compete with citrate for binding to Fe³⁺ and prevent the formation and adsorption of citrate-Fe³⁺ to the culture well. This technique was validated with the known iron chelators deferiprone and deferoxamine, and with the bacterial siderophore 2,3-dihydroxybenzoic acid and the catechol carbidopa.     

Iron Metabolism in Aging and Age-Related Diseases

Iron is a trace metal element necessary to maintain life and is also involved in a variety of biological processes. Aging refers to the natural life process in which the physiological functions of the various systems, organs, and tissues decline, affected by genetic and environmental factors. Therefore, it is imperative to investigate the relationship between iron metabolism and aging-related diseases, including neurodegenerative diseases. During aging, the accumulation of nonheme iron destroys the stability of the intracellular environment. The destruction of iron homeostasis can induce cell damage by producing hydroxyl free radicals, leading to mitochondrial dysfunction, brain aging, and even organismal aging. In this review, we have briefly summarized the role of the metabolic process of iron in the body, then discussed recent developments of iron metabolism in aging and age-related neurodegenerative diseases, and finally, explored some iron chelators as treatment strategies for those disorders. Understanding the roles of iron metabolism in aging and neurodegenerative diseases will fill the knowledge gap in the field. This review could provide new insights into the research on iron metabolism and age-related neurodegenerative diseases.     

Iron at the Interface of Hepatocellular Carcinoma

Cancer incidence and mortality are rapidly growing, with liver cancer being the sixth most diagnosed cancer worldwide and the third leading cause of cancer death in 2020. A number of risk factors have been identified that trigger the progression to hepatocellular carcinoma. In this review, we focus on iron as a potential risk factor for liver carcinogenesis. Molecules involved in the regulation of iron metabolism are often upregulated in cancer cells, in order to provide a supply of this essential trace element for all stages of tumor development, survival, proliferation, and metastasis. Thus, cellular and systemic iron levels must be tightly regulated to prevent or delay liver cancer progression. Disorders associated with dysregulated iron metabolism are characterized with increased susceptibility to hepatocellular carcinoma. This review discusses the association of iron with metabolic disorders such as hereditary hemochromatosis, non-alcoholic fatty liver disease, obesity, and type 2 diabetes, in the background of hepatocellular carcinoma.     

Protease and Amylase Stability in the Presence of Chelators Used in Laundry Detergent Applications: Correlation Between Chelator Properties and Enzyme Stability in Liquid Detergents

Chelators are a common ingredient in most laundry detergents. They have a number of different functions such as reducing water hardness, assisting in keeping particulate soil in suspension and the removal of certain stains, thus complementing the action of the anionic surfactants. Another important group of components in a modern liquid detergent is enzymes, mainly proteases and amylases. As the most commonly used enzymes within the detergent industry are dependent on bound calcium ions to maintain conformational stability and function, the presence of both chelators and enzymes in a liquid detergent presents a challenge. The three commonly used Ca²⁺ chelators: citrate, DTPA (diethylene triamine pentaacetic acid) and HEDP (1-hydroxyethane-1,1-diyl)bis(phosphonic acid), were studied with regard to their impact on protease and amylase stability in buffer and in a model liquid detergent. Enzyme stability was characterized by differential scanning calorimetry (DSC) and activity studies, and correlated to the chelator-Ca²⁺ interaction properties. The results show that a chelator’s ability to reduce water hardness and its Ca²⁺ affinity are in reality two separate aspects in the context of their use in liquid detergents. In the presence of DTPA, stoichiometric surplus of free Ca²⁺ is required to maintain sufficient amylase and protease stability. In the presence of the weaker chelators, HEDP and citrate, the total Ca²⁺ concentration is more important to protein stability than stoichiometric balancing between chelator and Ca²⁺. Thus, for these chelators their total concentration only has a minor impact on the Ca²⁺ concentration required to maintain or improve enzyme storage stability. The results underline the importance of Ca²⁺ in liquid detergent formulations, and suggest how proper balancing of chelators and Ca²⁺ can be used to improve overall enzyme stability.     

The influence of different chelators on the solubilization/liquefaction of different pretreated and natural lignites

The question was investigated, whether chelators alone can solubilize different lignites. For that purpose the influence of eight chelators were tested on different pretreated or natural lignites. The effects of the pretreatment and of the concentration of chelators were studied. The results show that different chelators have different influence on different lignites. Oxidative pretreatment enhances the solubilization. At low chelator concentrations only leonardite is solubilized to remarkable degree.

Ammonium oxalate is one of the most effective agents. The results confirm that chelators alone are not able to solubilize lignites with a very low ash or oxygen content.     

Redox cycling of iron and lipid peroxidation

Mechanisms of iron-catalyzed lipid peroxidation depend on the presence or absence of preformed lipid hydroperoxides (LOOH). Preformed LOOH are decomposed by Fe(II) to highly reactive lipid alkoxyl radicals, which in turn promote the formation of new LOOH. However, in the absence of LOOH, both Fe²⁺ and Fe³⁺ must be available to initiate lipid peroxidation, with optimum activity occurring as the Fe²⁺/Fe³⁺ ratio approaches unity. The simultaneous availability of Fe²⁺ and Fe³⁺ can be achieved by oxidizing some Fe²⁺ with hydrogen peroxide or with chelators that favor autoxidation of Fe²⁺ by molecular oxygen. Alternatively, one can use Fe³⁺ and reductants like superoxide, ascorbate or thiols. In either case excess Fe²⁺ oxidation or Fe³⁺ reduction will inhibit lipid peroxidation by converting all the iron to the Fe³⁺ or Fe²⁺ form, respectively. Superoxide dismutase and catalase can affect lipid peroxidation by affecting iron reduction/oxidation and the formation of a (1∶1) Fe²⁺/Fe³⁺ ratio. Hydroxyl radical scavengers can also increase or decrease lipid peroxidation by affecting the redox cycling of iron. Based on a paper presented at the Symposium on Metals and Lipid Oxidation, held at the AOCS Annual Meeting in Baltimore, Maryland, April 1990.     

The Iron Chelator Desferrioxamine Increases the Efficacy of Bedaquiline in Primary Human Macrophages Infected with BCG

For over 50 years, patients with drug-sensitive and drug-resistant tuberculosis have undergone long, arduous, and complex treatment processes with several antimicrobials. With the prevalence of drug-resistant strains on the rise and new therapies for tuberculosis urgently required, we assessed whether manipulating iron levels in macrophages infected with mycobacteria offered some insight into improving current antimicrobials that are used to treat drug-resistant tuberculosis. We investigated if the iron chelator, desferrioxamine, can support the function of human macrophages treated with an array of second-line antimicrobials, including moxifloxacin, bedaquiline, amikacin, clofazimine, linezolid and cycloserine. Primary human monocyte-derived macrophages were infected with Bacillus Calmette-Guérin (BCG), which is pyrazinamide-resistant, and concomitantly treated for 5 days with desferrioxamine in combination with each one of the second-line tuberculosis antimicrobials. Our data indicate that desferrioxamine used as an adjunctive treatment to bedaquiline significantly reduced the bacterial load in human macrophages infected with BCG. Our findings also reveal a link between enhanced bactericidal activity and increases in specific cytokines, as the addition of desferrioxamine increased levels of IFN-γ, IL-6, and IL-1β in BCG-infected human monocyte-derived macrophages (hMDMs) treated with bedaquiline. These results provide insight, and an in vitro proof-of-concept, that iron chelators may prove an effective adjunctive therapy in combination with current tuberculosis antimicrobials.     

Recent Advances in Hydrogen Peroxide Responsive Organoborons for Biological and Biomedical Applications

Hydrogen peroxide is the most stable reactive oxygen species generated endogenously, participating in numerous physiological processes and abnormal pathological conditions. Mounting evidence suggests that a higher level of H₂O₂ exists in various disease conditions. Thus, H₂O₂ functions as an ideal target for site-specific bioimaging and therapeutic targeting. The unique reactivity of organoborons with H₂O₂ provides a method for developing chemoselective molecules for biological and biomedical applications. This review highlights the design and application of boron-derived molecules for H₂O₂ detection, and the utility of boron moieties toward masking reactive compounds leading to the development of metal prochelators and prodrugs for selectively delivering an active species at the target sites with elevated H₂O₂ levels. Additionally, the emergence of H₂O₂-responsive theranostic agents consisting of both therapeutic and diagnostic moieties in one integrated system are discussed. The purpose of this review is to provide a better understanding of the role of boron-derived molecules toward biological and pharmacological applications.     

Approaching ‘Kit‐Type’ Labelling with 68Ga: The DATA Chelators

The DATA chelators are a novel class of tri‐anionic ligands based on 6‐amino‐1,4‐ d i a zepine‐ t riacetic a cid, which have been introduced recently for the chelation of ⁶⁸Ga. Compared with macrocyclic chelators based on the cyclen scaffold (i.e., DOTA, DO3A, and DO2A derivatives), DATA chelators undergo quantitative radiolabelling more rapidly and under milder conditions. In this study, a systematic evaluation of the labelling of four DATA chelators—DATA^M, DATA^P, DATA^Ph, and DATA^PPh—with ⁶⁸Ga is presented. The results highlight the extraordinary potential of this new class of chelators for application in molecular imaging using ⁶⁸Ga positron emission tomography (PET).     

A microsomal membrane component associated with iron reduction in NADPH-supported lipid peroxidation

This study was conducted to determine whether a factor responsible for reduced nicotinamide adenine dinucleotide phosphate (NADPH)-supported lipid peroxidation in rat liver microsomes is involved in iron reduction by cooperation with NADPH-cytochrome P450 reductase. Under anaerobic conditions, NADPH-dependent reduction of ferric pyrophosphate in microsomes was not dependent on cytochrome P450 levels and was not inhibited by carbon monoxide (CO). All of the iron complexes with chelators such as adenosine 5′-diphosphate, pyrophosphate, nitrilotriacetate, oxalate or citrate were reduced in microsomes, although in the reconstituted system containing purified NADPH-cytochrome P450 reductase little or no iron reduction was found. A cytochrome P450-free fraction from a cholate-solubilized preparation of microsomes after passage through a laurate sepharose column was required for reduction of iron pyrophosphate in the reconstituted system leading to lipid peroxidation. The iron reduction was not inhibited by CO and was destroyed by heat treatment or trypsin digestion of the fraction. All iron complexes were reduced in the presence of the fraction, using a reducing equivalent of NADPHvia NADPH-cytochrome P450 reductase. The results indicate that a heat-labile component, which is probably a protein distinct from cytochrome P450, is associated with iron reduction responsible for lipid peroxidation in microsomes.     

Inhibitors of the salicylate synthase (MbtI) from Mycobacterium tuberculosis discovered by high-throughput screening

A simple steady‐state kinetic high‐throughput assay was developed for the salicylate synthase MbtI from Mycobacterium tuberculosis, which catalyzes the first committed step of mycobactin biosynthesis. The mycobactins are small‐molecule iron chelators produced by M. tuberculosis, and their biosynthesis has been identified as a promising target for the development of new antitubercular agents. The assay was miniaturized to a 384‐well plate format and high‐throughput screening was performed at the National Screening Laboratory for the Regional Centers of Excellence in Biodefense and Emerging Infectious Diseases (NSRB). Three classes of compounds were identified comprising the benzisothiazolones (class I), diarylsulfones (class II), and benzimidazole‐2‐thiones (class III). Each of these compound series was further pursued to investigate their biochemical mechanism and structure–activity relationships. Benzimidazole‐2‐thione 4 emerged as the most promising inhibitor owing to its potent reversible inhibition.     

Why nutraceuticals do not prevent or treat Alzheimer's disease

A great deal of research has pointed to deleterious roles of metal ions in the development of Alzheimer's disease. These include: i) the precipitation and aggregation of amyloid β (Aβ) peptides to form senile plaques and neurofibrillary tangles, and/or ii) the augmentation of oxidative stress by metal ion mediated production and activation of hydrogen peroxide. The growing trend in nutraceutical intake is in part a result of the belief that they postpone the development of dementias such as Alzheimer's disease. However, pathogenic events centred on metal ions are expected to be aggravated by frequent nutraceutical intake. Novel therapeutic approaches centred on chelators with specificity for copper and iron ions should be fully explored.     

Scavenging Bacterial Siderophores with Engineered Lipocalin Proteins as an Alternative Antimicrobial Strategy

Iron acquisition mediated by siderophores, high-affinity chelators for which bacteria have evolved specific synthesis and uptake mechanisms, plays a crucial role in microbiology and in host–pathogen interactions. In the ongoing fight against bacterial infections, this area has attracted biomedical interest. Beyond several approaches to interfere with siderophore-mediated iron uptake from medicinal and immunochemistry, the development of high-affinity protein scavengers that tightly complex the siderophores produced by pathogenic bacteria has appeared as a novel strategy. Such binding proteins have been engineered based on siderocalin—also known as lipocalin 2—an endogenous human scavenger of enterobactin and bacillibactin that controls the systemic spreading of commensal bacteria such as Escherichia coli. By using combinatorial protein design, siderocalin was reshaped to bind several siderophores from Pseudomonas aeruginosa and, in particular, petrobactin from Bacillus anthracis, none of which is recognized by the natural protein. Such engineered versions of siderocalin effectively suppress the growth of corresponding pathogenic bacteria by depriving them of their iron supply and offer the potential to complement antibiotic therapy in situations of acute or persistent infection.