Tumor Cell Cytotoxicity of a Novel Metal Chelator

Abstract We have synthesized a novel six-coordinate metal chelator from the triamine cis-1,3,5-triaminocyclohexane by the addition of a 2-pyridylmethyl pendant arm on each nitrogen, which we term tachpyr. The experiments described here were designed to explore whether this compound exhibits potential antitumor activity. When added to MBT2 or T24 cultured bladder cancer cells, tachpyr was profoundly cytotoxic, with an IC50 of approximately 4.6 μmol/L compared with 70 μmol/L for desferioxamine. To explore the mode of action of tachpyr, several metal complexes were prepared, including Fe(II), Ca(II), Mn(II), Mg(II), Cu(II), and Zn(II) tachpyr complexes. Of these, the Zn(II), Cu(II), and Fe(II) complexes were without toxic effect, whereas the Ca(II), Mn(II), and Mg(II) complexes remained cytotoxic. To further probe the role of Zn(II) and Cu(II) chelation in the cytotoxicity of tachpyr, sterically hindered tachpyr derivatives were prepared through N-alkylation of tachpyr. These derivatives were unable to strongly bind Fe(III) or Fe(II) but were able to bind Zn(II) and Cu(II). When added to cells, these sterically hindered tachpyr derivatives were nontoxic, consistent with a role of iron depletion in the cytotoxic mechanism of tachpyr. Further, the addition of tachpyr to proliferating cultures resulted in an early and selective inhibition of ferritin synthesis, an iron storage protein whose translation is critically dependent on intracellular iron pools. Taken together, these experiments suggest that tachpyr is a cytotoxic metal chelator that targets intracellular iron, and that the use of tachpyr in cancer therapy deserves further exploration. © 1998 by The American Society of Hematology.

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Tumor Cell Cytotoxicity of a Novel Metal Chelator

Blood ◽

10.1182/blood.v92.4.1384.416k30_1384_1389 ◽

1998 ◽

Vol 92 (4) ◽

pp. 1384-1389 ◽

Cited By ~ 1

Author(s):

S.V. Torti ◽

F.M. Torti ◽

S.P. Whitman ◽

M.W. Brechbiel ◽

G. Park ◽

...

Keyword(s):

Selective Inhibition ◽

Intracellular Iron ◽

Iron Storage ◽

Metal Chelator ◽

Pendant Arm ◽

Ferritin Synthesis ◽

Bladder Cancer Cells ◽

Sterically Hindered ◽

American Society

We have synthesized a novel six-coordinate metal chelator from the triamine cis-1,3,5-triaminocyclohexane by the addition of a 2-pyridylmethyl pendant arm on each nitrogen, which we term tachpyr. The experiments described here were designed to explore whether this compound exhibits potential antitumor activity. When added to MBT2 or T24 cultured bladder cancer cells, tachpyr was profoundly cytotoxic, with an IC50 of approximately 4.6 μmol/L compared with 70 μmol/L for desferioxamine. To explore the mode of action of tachpyr, several metal complexes were prepared, including Fe(II), Ca(II), Mn(II), Mg(II), Cu(II), and Zn(II) tachpyr complexes. Of these, the Zn(II), Cu(II), and Fe(II) complexes were without toxic effect, whereas the Ca(II), Mn(II), and Mg(II) complexes remained cytotoxic. To further probe the role of Zn(II) and Cu(II) chelation in the cytotoxicity of tachpyr, sterically hindered tachpyr derivatives were prepared through N-alkylation of tachpyr. These derivatives were unable to strongly bind Fe(III) or Fe(II) but were able to bind Zn(II) and Cu(II). When added to cells, these sterically hindered tachpyr derivatives were nontoxic, consistent with a role of iron depletion in the cytotoxic mechanism of tachpyr. Further, the addition of tachpyr to proliferating cultures resulted in an early and selective inhibition of ferritin synthesis, an iron storage protein whose translation is critically dependent on intracellular iron pools. Taken together, these experiments suggest that tachpyr is a cytotoxic metal chelator that targets intracellular iron, and that the use of tachpyr in cancer therapy deserves further exploration. © 1998 by The American Society of Hematology.

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Nitric Oxide–Mediated Induction of Ferritin Synthesis in J774 Macrophages by Inflammatory Cytokines: Role of Selective Iron Regulatory Protein-2 Downregulation

Blood ◽

10.1182/blood.v91.3.1059.1059_1059_1066 ◽

1998 ◽

Vol 91 (3) ◽

pp. 1059-1066 ◽

Cited By ~ 1

Author(s):

Stefania Recalcati ◽

Donatella Taramelli ◽

Dario Conte ◽

Gaetano Cairo

Keyword(s):

Nitric Oxide ◽

Posttranscriptional Regulation ◽

Iron Homeostasis ◽

Regulatory Protein ◽

Intracellular Iron ◽

Iron Regulatory Proteins ◽

Ferritin Synthesis ◽

Direct Demonstration ◽

J774 Cells

Cytokine-treated macrophages represent a useful model to unravel the molecular basis of reticuloendothelial (RE) iron retention in inflammatory conditions. In the present study, we showed that stimulation of murine macrophage J774 cells with interferon (IFN)-γ/lipopolysaccharide (LPS) resulted in a nitric oxide-dependent modulation of the activity of iron regulatory proteins (IRP)-1 and 2, cytoplasmic proteins which, binding to RNA motifs called iron responsive elements (IRE), control ferritin translation. Stimulation with cytokines caused a small increase of IRP-1 activity and a strong reduction of IRP-2 activity accompanied by increased ferritin synthesis and accumulation. Cytokines induced only a minor increase of H chain ferritin mRNA, thus indicating that IRP-2–mediated posttranscriptional regulation plays a major role in the control of ferritin expression. This was confirmed by direct demonstration that the translational repression function of IRP was impaired in stimulated cells. In fact, translation in cell-free extracts of a reporter transcript under the control of an IRE sequence was repressed less efficiently by IRP-containing lysates from cytokine-treated cells than by lysates from control cells. Our findings throw light on the role of IRP-2 showing that: (1) this protein responds to a stimulus in opposite fashion to IRP-1; (2) when abundantly expressed, as in J774 cells, IRP-2 is sufficient to regulate intracellular iron metabolism in living cells; and (3) by allowing increased ferritin synthesis, IRP-2 may play a role in the regulation of iron homeostasis in RE cells during inflammation.

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HFE Downregulates Iron Uptake From Transferrin and Induces Iron-Regulatory Protein Activity in Stably Transfected Cells

Blood ◽

10.1182/blood.v94.11.3915.423k27_3915_3921 ◽

1999 ◽

Vol 94 (11) ◽

pp. 3915-3921 ◽

Cited By ~ 2

Author(s):

H.D. Riedel ◽

M.U. Muckenthaler ◽

S.G. Gehrke ◽

I. Mohr ◽

K. Brennan ◽

...

Keyword(s):

Transferrin Receptor ◽

Iron Uptake ◽

Iron Homeostasis ◽

Hereditary Hemochromatosis ◽

Regulatory Protein ◽

Intracellular Iron ◽

Iron Storage ◽

Iron Regulatory Proteins ◽

Cellular Iron

Hereditary hemochromatosis (HH) is a common autosomal-recessive disorder of iron metabolism. More than 80% of HH patients are homozygous for a point mutation in a major histocompatibility complex (MHC) class I type protein (HFE), which results in a lack of HFE expression on the cell surface. A previously identified interaction of HFE and the transferrin receptor suggests a possible regulatory role of HFE in cellular iron absorption. Using an HeLa cell line stably transfected with HFE under the control of a tetracycline-sensitive promoter, we investigated the effect of HFE expression on cellular iron uptake. We demonstrate that the overproduction of HFE results in decreased iron uptake from diferric transferrin. Moreover, HFE expression activates the key regulators of intracellular iron homeostasis, the iron-regulatory proteins (IRPs), implying that HFE can affect the intracellular “labile iron pool.” The increase in IRP activity is accompanied by the downregulation of the iron-storage protein, ferritin, and an upregulation of transferrin receptor levels. These findings are discussed in the context of the pathophysiology of HH and a possible role of iron-responsive element (IRE)-containing mRNAs.

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Synchrotron-Based Nano-XANES Reveals Intracellular Heterogeneity of Iron Species in Magnetotactic Bacteria

10.26434/chemrxiv.14461932.v1 ◽

2021 ◽

Author(s):

Daniel Chevrier ◽

Elisa Cerdá-Doñate ◽

Yeseul Park ◽

Fernando Cacho-Nerin ◽

Miguel Gomez-Gonzalez ◽

...

Keyword(s):

Cell Level ◽

Iron Species ◽

Intracellular Iron ◽

Iron Storage ◽

Edge Structure ◽

X Ray ◽

Magnetite Biomineralization ◽

Iron Material ◽

X Ray Absorption

<p>This report demonstrates how scanning X-ray fluorescence microscopy (SXFM) and nanoscale X-ray absorption near-edge structure (nano-XANES) can spatially and chemically identify intracellular iron species at the single-cell level, creating an opportunity to examine the role of iron storage in magnetite biomineralization. Fe K-edge nano-XANES measurements of <i>Magnetospirillum gryphiswaldense</i> in varied iron media conditions and iron storage capacity revealed intracellular iron heterogeneities through a distinction between formed magnetosomes and intracellular iron material. This work highlights the potential of nano-XANES in providing an experimental advantage in the multidisciplinary field of biomineralization.</p>

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Bacterioferritin: a key iron storage modulator that affects strain growth and butenyl-spinosyn biosynthesis in Saccharopolyspora pogona

Microbial Cell Factories ◽

10.1186/s12934-021-01651-x ◽

2021 ◽

Vol 20 (1) ◽

Author(s):

Jianli Tang ◽

Zirong Zhu ◽

Haocheng He ◽

Zhudong Liu ◽

Ziyuan Xia ◽

...

Keyword(s):

Stress Resistance ◽

Quantitative Proteomics ◽

Polyketide Synthase ◽

Storage Capacity ◽

Oxidative Stress Response ◽

Iron Toxicity ◽

Intracellular Iron ◽

Proteomics Analysis ◽

Iron Storage

Abstract Background Butenyl-spinosyn, produced by Saccharopolyspora pogona, is a promising biopesticide due to excellent insecticidal activity and broad pesticidal spectrum. Bacterioferritin (Bfr, encoded by bfr) regulates the storage and utilization of iron, which is essential for the growth and metabolism of microorganisms. However, the effect of Bfr on the growth and butenyl-spinosyn biosynthesis in S. pogona has not been explored. Results Here, we found that the storage of intracellular iron influenced butenyl-spinosyn biosynthesis and the stress resistance of S. pogona, which was regulated by Bfr. The overexpression of bfr increased the production of butenyl-spinosyn by 3.14-fold and enhanced the tolerance of S. pogona to iron toxicity and oxidative damage, while the knockout of bfr had the opposite effects. Based on the quantitative proteomics analysis and experimental verification, the inner mechanism of these phenomena was explored. Overexpression of bfr enhanced the iron storage capacity of the strain, which activated polyketide synthase genes and enhanced the supply of acyl-CoA precursors to improve butenyl-spinosyn biosynthesis. In addition, it induced the oxidative stress response to improve the stress resistance of S. pogona. Conclusion Our work reveals the role of Bfr in increasing the yield of butenyl-spinosyn and enhancing the stress resistance of S. pogona, and provides insights into its enhancement on secondary metabolism, which provides a reference for optimizing the production of secondary metabolites in actinomycetes.

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Differential Role of Ferritins in Iron Metabolism and Virulence of the Plant-Pathogenic Bacterium Erwinia chrysanthemi 3937

Journal of Bacteriology ◽

10.1128/jb.01640-07 ◽

2007 ◽

Vol 190 (5) ◽

pp. 1518-1530 ◽

Cited By ~ 35

Author(s):

Aïda Boughammoura ◽

Berthold F. Matzanke ◽

Lars Böttger ◽

Sylvie Reverchon ◽

Emmanuel Lesuisse ◽

...

Keyword(s):

Gene Expression ◽

Iron Homeostasis ◽

Reverse Genetics ◽

Bacterial Species ◽

Erwinia Chrysanthemi ◽

Intermediate Phenotype ◽

Redox Stress ◽

Intracellular Iron ◽

Iron Storage

ABSTRACT During infection, the phytopathogenic enterobacterium Erwinia chrysanthemi has to cope with iron-limiting conditions and the production of reactive oxygen species by plant cells. Previous studies have shown that a tight control of the bacterial intracellular iron content is necessary for full virulence. The E. chrysanthemi genome possesses two loci that could be devoted to iron storage: the bfr gene, encoding a heme-containing bacterioferritin, and the ftnA gene, coding for a paradigmatic ferritin. To assess the role of these proteins in the physiology of this pathogen, we constructed ferritin-deficient mutants by reverse genetics. Unlike the bfr mutant, the ftnA mutant had increased sensitivity to iron deficiency and to redox stress conditions. Interestingly, the bfr ftnA mutant displayed an intermediate phenotype for sensitivity to these stresses. Whole-cell analysis by Mössbauer spectroscopy showed that the main iron storage protein is FtnA and that there is an increase in the ferrous iron/ferric iron ratio in the ftnA and bfr ftnA mutants. We found that ftnA gene expression is positively controlled by iron and the transcriptional repressor Fur via the small antisense RNA RyhB. bfr gene expression is induced at the stationary phase of growth. The σS transcriptional factor is necessary for this control. Pathogenicity tests showed that FtnA and the Bfr contribute differentially to the virulence of E. chrysanthemi depending on the host, indicating the importance of a perfect control of iron homeostasis in this bacterial species during infection.

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The Role of Iron in the Pathogenesis of Atherosclerosis

Physiological Research ◽

10.33549/physiolres.933589 ◽

2017 ◽

pp. S55-S67 ◽

Cited By ~ 29

Author(s):

P. KRAML

Keyword(s):

Cardiovascular Risk ◽

Free Oxygen Radicals ◽

Intracellular Iron ◽

Iron Storage ◽

Oxidation Stress ◽

Iron Stores ◽

Labile Iron Pool ◽

Labile Iron ◽

Iron Pool

Ferritin and increased iron stores first appeared on the list of cardiovascular risk factors more than 30 years ago and their causal role in the pathogenesis of atherosclerosis has been heavily discussed since the early 1990s. It seems that besides traditional factors such as hyperlipoproteinemia, hypertension, diabetes mellitus, obesity, physical inactivity, smoking and family history, high iron stores represent an additional parameter that could modify individual cardiovascular risk. The role of iron in the pathogenesis of atherosclerosis was originally primarily associated with its ability to catalyze the formation of highly reactive free oxygen radicals and the oxidation of atherogenic lipoproteins. Later, it became clear that the mechanism is more complex. Atherosclerosis is a chronic fibroproliferative inflammatory process and iron, through increased oxidation stress as well as directly, can control both native and adaptive immune responses. Within the arterial wall, iron affects all of the cell types that participate in the atherosclerotic process (monocytes/macrophages, endothelial cells, vascular smooth muscle cells and platelets). Most intracellular iron is bound in ferritin, whereas redox-active iron forms labile iron pool. Pro-inflammatory and anti-inflammatory macrophages within arterial plaque differ with regard to the amount of intracellular iron and most probably with regard to their labile iron pool. Yet, the relation between plasma ferritin and intracellular labile iron pool has not been fully clarified. Data from population studies document that the consumption of meat and lack of physical activity contribute to increased iron stores. Patients with hereditary hemochromatosis, despite extreme iron storage, do not show increased manifestation of atherosclerosis probably due to the low expression of hepcidin in macrophages.

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Nitric Oxide–Mediated Induction of Ferritin Synthesis in J774 Macrophages by Inflammatory Cytokines: Role of Selective Iron Regulatory Protein-2 Downregulation

Blood ◽

10.1182/blood.v91.3.1059 ◽

1998 ◽

Vol 91 (3) ◽

pp. 1059-1066 ◽

Cited By ~ 101

Author(s):

Stefania Recalcati ◽

Donatella Taramelli ◽

Dario Conte ◽

Gaetano Cairo

Keyword(s):

Nitric Oxide ◽

Posttranscriptional Regulation ◽

Iron Homeostasis ◽

Regulatory Protein ◽

Intracellular Iron ◽

Iron Regulatory Proteins ◽

Ferritin Synthesis ◽

Direct Demonstration ◽

J774 Cells

Abstract Cytokine-treated macrophages represent a useful model to unravel the molecular basis of reticuloendothelial (RE) iron retention in inflammatory conditions. In the present study, we showed that stimulation of murine macrophage J774 cells with interferon (IFN)-γ/lipopolysaccharide (LPS) resulted in a nitric oxide-dependent modulation of the activity of iron regulatory proteins (IRP)-1 and 2, cytoplasmic proteins which, binding to RNA motifs called iron responsive elements (IRE), control ferritin translation. Stimulation with cytokines caused a small increase of IRP-1 activity and a strong reduction of IRP-2 activity accompanied by increased ferritin synthesis and accumulation. Cytokines induced only a minor increase of H chain ferritin mRNA, thus indicating that IRP-2–mediated posttranscriptional regulation plays a major role in the control of ferritin expression. This was confirmed by direct demonstration that the translational repression function of IRP was impaired in stimulated cells. In fact, translation in cell-free extracts of a reporter transcript under the control of an IRE sequence was repressed less efficiently by IRP-containing lysates from cytokine-treated cells than by lysates from control cells. Our findings throw light on the role of IRP-2 showing that: (1) this protein responds to a stimulus in opposite fashion to IRP-1; (2) when abundantly expressed, as in J774 cells, IRP-2 is sufficient to regulate intracellular iron metabolism in living cells; and (3) by allowing increased ferritin synthesis, IRP-2 may play a role in the regulation of iron homeostasis in RE cells during inflammation.

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The Role of NCOA4-Mediated Ferritinophagy in Iron Homeostasis

10.20944/preprints201809.0148.v1 ◽

2018 ◽

Cited By ~ 2

Author(s):

Naiara Santana-Codina ◽

Joseph D. Mancias

Keyword(s):

Iron Homeostasis ◽

Iron Release ◽

Intracellular Iron ◽

Nuclear Receptor Coactivator ◽

Iron Storage ◽

Physiological Processes ◽

Ferritin Iron ◽

Autophagic Degradation ◽

Dependent Cell

Nuclear receptor coactivator 4 (NCOA4) is a selective cargo receptor that mediates the autophagic degradation of ferritin (“ferritinophagy”), the cytosolic iron storage complex. NCOA4-mediated ferritinophagy maintains intracellular iron homeostasis by facilitating ferritin iron storage or release according to demand. Ferritinophagy is involved in iron-dependent physiological processes such as erythropoiesis, where NCOA4 mediates ferritin iron release for mitochondrial heme synthesis. Recently, ferritinophagy has been shown to regulate ferroptosis, a newly described form of iron-dependent cell death mediated by excess lipid peroxidation. Dysregulation of iron metabolism and ferroptosis have been described in neurodegeneration, cancer, and infection, but little is known about the role of ferritinophagy in the pathogenesis of these diseases. Here, we will review the biochemical regulation of NCOA4, its contribution to physiological processes and its role in disease. Finally, we will discuss the potential of activating or inhibiting ferritinophagy and ferroptosis for therapeutic purposes.

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