%0 Journal Article %~ PubMed %A Stefani, Christian %A Jansson, Patric J %A Gutierrez, Elaine %A Bernhardt, Paul V %A Richardson, Des R %A Kalinowski, Danuta S %T Alkyl Substituted 2'-Benzoylpyridine Thiosemicarbazone Chelators with Potent and Selective Anti-Neoplastic Activity: Novel Ligands that Limit Methemoglobin Formation. %B Journal of Medicinal Chemistry %D 2013 %C United States %I American Chemical Society %V 56 %N 1 %P 357-370 %@ 0022-2623 %X %Z FOR Codes: 111205 %0 Journal Article %~ PubMed %A Dixon, K M %A Lui, G Y L %A Kovacevic, Z %A Zhang, D %A Yao, M %A Chen, Z %A Dong, Q %A Assinder, S J %A Richardson, D R %T Dp44mT targets the AKT, TGF-β and ERK pathways via the metastasis suppressor NDRG1 in normal prostate epithelial cells and prostate cancer cells. %B British Journal of Cancer %D 2013 %C United Kingdom %I Nature Publishing Group %V 108 %N 2 %P 409-419 %@ 0007-0920 %X %Z FOR Codes: 60111 110199 %0 Journal Article %~ PubMed %A Bambang, I F %A Lee, Y K %A Richardson, D R %A Zhang, D %T Endoplasmic reticulum protein 29 regulates epithelial cell integrity during the mesenchymal-epithelial transition in breast cancer cells. %B Oncogene %D 2013 %C United Kingdom %I Nature Publishing Group %V 32 %N 10 %P 1240-1251 %@ 0950-9232 %X The epithelial-mesenchymal transition (EMT) correlates with disruption of cell-cell adhesion, loss of cell polarity and development of epithelial cell malignancy. Identifying novel molecules that inhibit EMT has profound potential for developing mechanism-based therapeutics. We previously demonstrated that the endoplasmic reticulum protein 29 (ERp29) is a novel factor that can drive mesenchymal-epithelial transition (MET) and induce cell growth arrest in MDA-MB-231 cells. Here, we show that ERp29 is an important molecule in establishing epithelial cell integrity during the MET. We demonstrate that ERp29 regulates MET in a cell context-dependent manner. ERp29 overexpression induced a complete MET in mesenchymal MDA-MB-231 cells through downregulating the expression of transcriptional repressors (for example, Slug, Snai1, ZEB2 and Twist) of E-cadherin. In contrast, overexpression of ERp29 induces incomplete MET in basal-like BT549 cells in which the expression of EMT-related markers (for example, vimentin; cytokeratin 19 (CK19) and E-cadherin) and the transcriptional repressors of E-cadherin were not altered. However, ERp29 overexpression in both cell-types resulted in loss of filamentous stress fibers, formation of cortical actin and restoration of an epithelial phenotype. Mechanistic studies revealed that overexpression of ERp29 in both cell-types upregulated the expression of TJ proteins (zonula-occludens-1 (ZO-1) and occludin) and the core apical-basal polarity proteins (Par3 and Scribble) at the membrane to enhance cell-cell contact and cell polarization. Knockdown of ERp29 in the epithelial MCF-7 cells decreased the expression of these proteins, leading to the disruption of cell-cell adhesion. Taken together, ERp29 is a novel molecule that regulates MET and epithelial cell integrity in breast cancer cells.Oncogene advance online publication, 30 April 2012; doi:10.1038/onc.2012.149. %Z FOR Codes: 111201 %0 Journal Article %~ PubMed %A Merlot, Angelica M %A Kalinowski, Danuta S %A Richardson, Des R %T Novel Chelators for Cancer Treatment: Where Are We Now? %B Antioxidants & Redox Signaling %D 2013 %C United States %I Mary Ann Liebert, Inc. Publishers %V 18 %N 8 %P 973-1006 %@ 1557-7716 %X Abstract Significance: Under normal circumstances, cellular iron levels are tightly regulated due to the potential toxic effects of this metal ion. There is evidence that tumors possess altered iron homeostasis, which is mediated by the perturbed expression of iron-related proteins, for example, transferrin receptor 1, ferritin and ferroportin 1. The de-regulation of iron homeostasis in cancer cells reveals a particular vulnerability to iron-depletion using iron chelators. In this review, we examine the absorption of iron from the gut; its transport, metabolism, and homeostasis in mammals; and the molecular pathways involved. Additionally, evidence for alterations in iron processing in cancer are described along with the perturbations in other biologically important transition metal ions, for example, copper(II) and zinc(II). These changes can be therapeutically manipulated by the use of novel chelators that have recently been shown to be highly effective in terms of inhibiting tumor growth. Recent Advances: Such chelators include those of the thiosemicarbazone class that were originally thought to target only ribonucleotide reductase, but are now known to have multiple effects, including the generation of cytotoxic radicals. Critical Issues: Several chelators have shown marked anti-tumor activity in vivo against a variety of solid tumors. An important aspect is the toxicology and the efficacy of these agents in clinical trials. Future Directions: As part of the process of the clinical assessment of the new chelators, an extensive toxicological assessment in multiple animal models is essential for designing appropriate dosing protocols in humans. Antioxid. Redox Signal. 00, 000-000. %Z FOR Codes: 111201 %0 Journal Article %~ PubMed %A Lukmantara, Adeline Y %A Kalinowski, Danuta S %A Kumar, Naresh %A Richardson, Des R %T Synthesis and biological evaluation of substituted 2-benzoylpyridine thiosemicarbazones: Novel structure-activity relationships underpinning their anti-proliferative and chelation efficacy. %B Bioorganic & Medicinal Chemistry Letters %D 2013 %C United Kingdom %I Pergamon %V 23 %N 4 %P 967-974 %@ 1464-3405 %X %Z FOR Codes: 111205 %0 Journal Article %~ PubMed %A Sun, Jing %A Zhang, Daohai %A Zheng, Ying %A Zhao, Qian %A Zheng, Minhua %A Kovacevic, Zaklina %A Richardson, Des R %T Targeting the Metastasis Suppressor, NDRG1, Using Novel Iron Chelators: Regulation of Stress Fiber-Mediated Tumor Cell Migration via Modulation of the ROCK1/pMLC2 Signaling Pathway. %B Molecular Pharmacology %D 2013 %C United States %I American Society for Pharmacology and Experimental Therapeutics %V 83 %N 2 %P 454-469 %@ 1521-0111 %X %Z FOR Codes: 111201 111501 %0 Journal Article %~ PubMed %A Lui, Goldie Y L %A Obeidy, Peyman %A Ford, Samuel J %A Tselepis, Chris %A Sharp, Danae M %A Jansson, Patric J %A Kalinowski, Danuta S %A Kovacevic, Zaklina %A Lovejoy, David B %A Richardson, Des R %T The Iron Chelator, Deferasirox, as a Novel Strategy for Cancer Treatment: Oral Activity Against Human Lung Tumor Xenografts and Molecular Mechanism of Action. %B Molecular Pharmacology %D 2013 %C United States %I American Society for Pharmacology and Experimental Therapeutics %V 83 %N 1 %P 179-190 %@ 1521-0111 %X %Z FOR Codes: 111205 %0 Journal Article %~ PubMed %A Kovacevic, Zaklina %A Chikhani, Sherin %A Lui, Goldie Y L %A Sivagurunathan, Sutharshani %A Richardson, Des R %T The Iron-Regulated Metastasis Suppressor NDRG1 Targets NEDD4L, PTEN, and SMAD4 and Inhibits the PI3K and Ras Signaling Pathways. %B Antioxidants & Redox Signaling %D 2013 %C United States %I Mary Ann Liebert, Inc. Publishers %V 18 %N 8 %P 874-887 %@ 1557-7716 %X Abstract Aims: The metastasis suppressor gene, N-myc downstream regulated gene-1 (NDRG1), is negatively correlated with tumor progression in multiple neoplasms, including pancreatic cancer. Moreover, NDRG1 is an iron-regulated gene that is markedly upregulated by cellular iron-depletion using novel antitumor agents such as the chelator, di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT), in pancreatic cancer cells. However, the exact function(s) of NDRG1 remain to be established and are important to elucidate. Results: In the current study, using gene-array analysis along with NDRG1 overexpression and silencing, we identified the molecular targets of NDRG1 in three pancreatic cancer cell lines. We demonstrate that NDRG1 upregulates neural precursor cell expressed developmentally downregulated 4-like (NEDD4L) and GLI-similar-3 (GLIS3). Further studies examining the downstream effects of NEDD4L led to the discovery that NDRG1 affects the transforming growth factor-?? (TGF-??) pathway, leading to the upregulation of two key tumor suppressor proteins, namely phosphatase and tensin homolog deleted on chromosome 10 (PTEN) and mothers against decapentaplegic homolog-4 (SMAD4). Moreover, NDRG1 inhibited the phosphatidylinositol 3-kinase (PI3K) and Ras oncogenic pathways. Innovation: This study provides significant insights into the mechanisms underlying the antitumor activity of NDRG1. For the first time, a role for NDRG1 is established in regulating the key signaling pathways involved in oncogenesis (TGF-??, PI3K, and Ras pathways). Conclusion: The identified target genes of NDRG1 and their effect on the TGF-?? signaling pathway reveal its molecular function in pancreatic cancer and a novel therapeutic avenue. Antioxid. Redox Signal. 00, 000-000. %Z FOR Codes: 1112 %0 Journal Article %~ PubMed %A Yu, Y %A Rahmanto, Y Suryo %A Richardson, Dr %T Bp44mT: An Orally-Active Iron Chelator of the Thiosemicarbazone Class with Potent Anti-Tumour Efficacy. %B British journal of pharmacology %D 2012 %C United Kingdom %I John Wiley & Sons Ltd. %V 165 %N 1 %P 148-66 %@ 0007-1188 %X Our previous studies demonstrated that a thiosemicarbazone iron chelator (di-2-pyridylketone-4,4-dimethyl-3-thiosemicarbazone; Dp44mT) possesses potent and selective anti-cancer activity but led to cardiotoxicity at non-optimal doses. In this study, we examined the in vivo anti-tumour efficacy and tolerability of a new-generation 2-benzoylpyridine thiosemicarbazone iron chelator (2-benzoylpyridine-4,4-dimethyl-3-thiosemicarbazone; Bp44mT) administered via the oral or i.v. routes. %Z FOR Codes: 111501 111204 %0 Journal Article %~ PubMed %A Debebe, Zufan %A Nekhai, Sergei %A Ashenafi, Meseret %A Lovejoy, David B %A Kalinowski, Danuta S %A Gordeuk, Victor R %A Byrnes, W Malcolm %A Richardson, Des R %A Karla, Pradeep K %T Development of a sensitive HPLC method to measure in vitro permeability of E- and Z-isomeric forms of thiosemicarbazones in Caco-2 monolayers. %B Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences %D 2012 %C Netherlands %I Elsevier BV %V 906 %N %P 25-32 %@ 1873-376X %X %Z FOR Codes: 111205 %0 Journal Article %~ PubMed %A Gao, Danmei %A Bambang, I Fon %A Putti, Thomas C %A Lee, Yuan Kun %A Richardson, Des R %A Zhang, Daohai %T ERp29 induces breast cancer cell growth arrest and survival through modulation of activation of p38 and upregulation of ER stress protein p58(IPK). %B Laboratory Investigation %D 2012 %C United States %I Nature Publishing Group %V 92 %N 2 %P 200-213 %@ 1530-0307 %X Endoplasmic reticulum protein 29 (ERp29) is an ER luminal protein that has a role in protein unfolding and secretion, but its role in cancer is unclear. Recently, we reported that overexpression of ERp29 significantly inhibited cell proliferation and prevented tumorigenesis in highly proliferative MDA-MB-231 breast cancer cells. Here, we show that ERp29-induced cancer cell growth arrest is modulated by the interplay between the concomitant phosphorylation of p38 and upregulation of the inhibitor of the interferon-induced, double-stranded RNA-activated protein kinase, p58(IPK). In this cell model, ERp29 overexpression significantly downregulates modulators of cell proliferation, namely urokinase plasminogen activator receptor, ?(1)-integrin and epidermal growth factor receptor. Furthermore, ERp29 significantly (P<0.001) increases phosphorylation of p38 (p-p38) and reduces matrix metalloproteinase-9 secretion. The role of ERp29 in upregulating cyclin-dependent kinase inhibitors (p15 and p21) and in downregulating cyclin D(2) is demonstrated in slowly proliferating ERp29-overexpressing MDA-MB-231 cells, whereas the opposite response was observed in ERp29-knockdown MCF-7 cells. Pharmacological inhibition of p-p38 downregulates p15 and p21 and inhibits eIF2? phosphorylation, indicating a role for p-p38 in this process. Furthermore, p58(IPK) expression was increased in ERp29-overexpressing MDA-MB-231 cells and highly decreased in ERp29-knockdown MCF-7 cells. This upregulation of p58(IPK) by ERp29 suppresses the activation of p-p38/p-PERK/p-eIF2? by repressing eIF2? phosphorylation. In fact, reduction of p58(IPK) expression by RNA interference stimulated eIF2? phosphorylation. The repression of eIF2? phosphorylation by p58(IPK) prevents ERp29-transfected cells from undergoing ER-dependent apoptosis driven by the activation of ATF4/CHOP/caspase-3. Hence, the interplay between p38 phosphorylation and p58(IPK) upregulation has key roles in modulating ERp29-induced cell-growth arrest and survival. %Z FOR Codes: 60111 %0 Journal Article %~ PubMed %A Whitnall, Megan %A Rahmanto, Yohan Suryo %A Huang, Michael L-H %A Saletta, Federica %A Lok, Hiu Chuen %A Gutiérrez, Lucía %A Lázaro, Francisco J %A Fleming, Adam J %A St Pierre, Tim G %A Mikhael, Marc R %A Ponka, Prem %A Richardson, Des R %T Identification of nonferritin mitochondrial iron deposits in a mouse model of Friedreich ataxia. %B Proceedings of the National Academy of Sciences %D 2012 %C United States %I National Academy of Sciences %V 109 %N 50 %P 20590-20595 %@ 0027-8424 %X %Z FOR Codes: 601 %0 Journal Article %~ PubMed %A Brychtova, Katerina %A Dvorakova, Lenka %A Opatrilova, Radka %A Raich, Ivan %A Kacerova, Sandra %A Placek, Lukas %A Kalinowski, Danuta S %A Richardson, Des R %A Jampilek, Josef %T Investigation of substituted 6-aminohexanoates as skin penetration enhancers. %B Bioorganic & Medicinal Chemistry %D 2012 %C United Kingdom %I Pergamon %V 20 %N 1 %P 86-95 %@ 0968-0896 %X Skin penetration enhancers are compounds used to facilitate the transdermal delivery of drugs that are otherwise not sufficiently permeable. Through a synthetic route implementing two series of esters, we generated transdermal penetration enhancers by a multi-step reaction with substituted 6-aminohexanoic acid. We present the synthesis of all newly prepared compounds here with structural confirmation accomplished by (1)H NMR, (13)C NMR, IR and mass spectroscopy (MS). The lipophilicity (logk) of all compounds was determined via RP-HPLC and their hydrophobicity (logP/ClogP) was also calculated using two commercially available programs. Ab initio calculations of geometry and molecular dynamic simulations were employed to investigate the 3-dimensional structures of selected compounds. The transdermal penetration-enhancing activity of all the synthesized esters were examined in vitro and demonstrated higher enhancement ratios than oleic acid. Compounds 2e (C(10) ester chain) and 2f (C(11) ester chain) exhibited the highest enhancement ratios. It can be concluded that the series non-substituted at the C((2)) position by a ?-lactam ring showed significantly higher activity than those with azepan-2-one. None of the prepared compounds penetrated through the skin. All of the investigated agents demonstrated minimal anti-proliferative activity using the SK-N-MC neuroepithelioma cell line (IC(50)>6.25?M), suggesting these analogs would have a low cytotoxic profile when administered in vivo as chemical penetration enhancers. The correlation between the chemical structure of the studied compounds and their lipophilicity is discussed in regards to transdermal penetration-enhancing activity. %Z FOR Codes: 111504 %0 Journal Article %~ PubMed %A Yu, Y %A Gutierrez, E %A Kovacevic, Z %A Saletta, F %A Obeidy, P %A Rahmanto, Y Suryo %A Richardson, D R %T Iron chelators for the treatment of cancer. %B Current Medicinal Chemistry %D 2012 %C Netherlands %I Bentham Science Publishers Ltd. %V 19 %N 17 %P 2689-2702 %@ 1875-533X %X The study of iron chelators as anti-tumor agents is still in its infancy. Iron is important for cellular proliferation and this is demonstrated by observations that iron-depletion results in cell cycle arrest and also apoptosis. In addition, many iron chelators are known to inhibit ribonucleotide reductase, the iron-containing enzyme that is the rate-limiting step for DNA synthesis. Desferrioxamine is a well known chelator used for the treatment of iron-overload disease, but it has also been shown to possess anti-cancer activity. Another class of chelators, namely the thiosemicarbazones, have been shown to possess anti-cancer activity since the 1950''s, although their mechanism(s) of action have only recently been more comprehensively elucidated. In fact, the redox activity of thiosemicarbazone iron complexes is thought to be important in mediating their potent cytotoxicity. Moreover, unlike typical iron chelators which simply act to deplete tumors of iron, several thiosemicarbazones (i.e., Bp44mT and Dp44mT) do not induce this effect, their anti-cancer efficacy being due to other mechanisms e.g., redox activity. Other reports have also shown that some thiosemicarbazones inhibit topoisomerase II??, demonstrating that this class of agents have multiple molecular targets and act by various mechanisms. The most well characterized thiosemicarbazone iron chelator in terms of its assessment in humans is 3-aminopyridine-2-carboxaldehyde thiosemicarbazone (3-AP). Observations from these clinical trials highlight the less than optimal activity of this ligand and several side effects related to its use, including myelo-suppression, hypoxia and methemoglobinemia. The mechanisms responsible for these latter effects must be elucidated and the design of the ligand altered to minimize these problems and increase efficacy. This review discusses the development of chelators as unique agents for cancer treatment. %Z FOR Codes: 1112 %0 Journal Article %~ PubMed %A Stariat, Jan %A Sestak, Vit %A Vavrova, Katerina %A Nobilis, Milan %A Kollarova, Zuzana %A Klimes, Jiri %A Kalinowski, Danuta S %A Richardson, Des R %A Kovarikova, Petra %T LC-MS/MS identification of the principal in vitro and in vivo phase I metabolites of the novel thiosemicarbazone anti-cancer drug, Bp4eT. %B Analytical and bioanalytical chemistry %D 2012 %C Germany %I Springer %V 403 %N 1 %P 309-321 %@ 1618-2650 %X %Z FOR Codes: 110101 111201 %0 Journal Article %~ PubMed %A Suryo Rahmanto, Yohan %A Bal, Sumeet %A Loh, Kim H %A Yu, Yu %A Richardson, Des R %T Melanotransferrin: Search for a function. %B Biochimica et Biophysica Acta %D 2012 %C Netherlands %I Elsevier BV %V 1820 %N 3 %P 237-243 %@ 0006-3002 %X Melanotransferrin was discovered in the 1980s as one of the first melanoma tumour antigens. The molecule is a transferrin homologue that is found predominantly bound to the cell membrane by a glycosyl-phosphatidylinositol anchor. MTf was described as an oncofoetal antigen expressed in only small quantities in normal tissues, but in much larger amounts in neoplastic cells. Several diseases are associated with expression of melanotransferrin, including melanoma and Alzheimer''s disease, although the significance of the protein to the pathogenesis of these conditions remains unclear. %Z FOR Codes: 111207 60103 %0 Journal Article %~ PubMed %A Quach, Patricia %A Gutierrez, Elaine %A Basha, Maram Talal %A Kalinowski, Danuta S %A Sharpe, Philip C %A Lovejoy, David B %A Bernhardt, Paul V %A Jansson, Patric J %A Richardson, Des R %T Methemoglobin Formation by 3-AP, Dp44mT and Other Anti-Cancer Thiosemicarbazones: Identification of Novel Thiosemicarbazones and Therapeutics that Prevent this Effect. %B Molecular Pharmacology %D 2012 %C United States %I American Society for Pharmacology and Experimental Therapeutics %V 82 %N 1 %P 105-114 %@ 1521-0111 %X Thiosemicarbazones are a group of compounds that have received comprehensive investigation as anti-cancer agents. Recently, the anti-tumor activity of the thiosemicarbazone, 3-amino-2-pyridinecarboxaldehyde thiosemicarbazone (3-AP, Triapine(??)), has been extensively assessed in over 20 Phase I and II clinical trials. These studies have demonstrated that 3-AP induces methemoglobin (metHb)-formation and hypoxia in patients, limiting its usefulness. Considering this problem, we assessed the mechanism of metHb-formation by 3-AP compared to more recently developed thiosemicarbazones, including di-2-pyridylketone-4,4-dimethyl-3-thiosemicarbazone (Dp44mT). This was investigated using intact red blood cells (RBCs), RBC lysates, purified oxyhemoglobin and a mouse model. The chelation of cellular labile iron with the formation of a redox-active thiosemicarbazone-iron complex was found to be crucial for oxyhemoglobin oxidation. This observation was substantiated using a thiosemicarbazone that cannot ligate iron and also by using the chelator, desferrioxamine (DFO), that forms a redox-inactive iron complex. Significantly, cellular copper chelation was not important for metHb generation in contrast to its role in preventing tumor cell proliferation. Administration of Dp44mT to mice catalyzed metHb- and cardiac metmyoglobin formation. However, ascorbic acid (AA) administered together with the drug in-vivo significantly decreased metHb levels, providing a potential therapeutic intervention. Moreover, we demonstrated that the structure of the thiosemicarbazone is of importance in terms of metHb generation, as the DpT analog, di-2-pyridylketone-4-cyclohexyl-4-methyl-3-thiosemicarbazone (DpC), does not induce metHb-generation in-vivo. Hence, DpC represents a next generation thiosemicarbazone that possesses markedly superior properties. This investigation is important for developing more effective thiosemicarbazone treatment regimens. %Z FOR Codes: 111506 %0 Journal Article %~ PubMed %A Lok, Hiu Chuen %A Suryo Rahmanto, Yohan %A Hawkins, Clare L %A Kalinowski, Danuta S %A Morrow, Charles S %A Townsend, Alan J %A Ponka, Prem %A Richardson, Des R %T Nitric oxide storage and transport in cells is mediated by glutathione-S-transferase P1-1 and Multidrug Resistant Protein 1 via Dinitrosyl iron complexes. %B Journal of Biological Chemistry %D 2012 %C United States %I American Society for Biochemistry and Molecular Bi %V 287 %N 1 %P 607-618 %@ 0021-9258 %X Nitrogen monoxide (NO) plays a role in the cytotoxic mechanisms of activated macrophages against tumor cells by inducing iron release. We showed that NO-mediated iron efflux from cells required glutathione (GSH) (Watts, R. N., and Richardson, D. R. (2001) J. Biol. Chem. 276, 4724-4732) and that the GSH-conjugate transporter, multidrug resistance-associated protein 1 (MRP1), mediates this release potentially as a dinitrosyl-dithiol iron complex (DNIC; Watts, R. N., Hawkins, C., Ponka, P., and Richardson, D. R. (2006) Proc. Natl. Acad. Sci. U.S.A. 103, 7670-7675). Recently, glutathione S-transferase P1-1 (GST P1-1) was shown to bind DNICs as dinitrosyl-diglutathionyl iron complexes. Considering this and that GSTs and MRP1 form an integrated detoxification unit with chemotherapeutics, we assessed whether these proteins coordinately regulate storage and transport of DNICs as long lived NO intermediates. Cells transfected with GSTP1 (but not GSTA1 or GSTM1) significantly decreased NO-mediated 59Fe release from cells. This NO-mediated 59Fe efflux and the effect of GST P1-1 on preventing this were observed with NO-generating agents and also in cells transfected with inducible nitric oxide synthase. Notably, 59Fe accumulated in cells within GST P1-1-containing fractions, indicating an alteration in intracellular 59Fe distribution. Furthermore, electron paramagnetic resonance studies showed that MCF7-VP cells transfected with GSTP1 contain significantly greater levels of a unique DNIC signal. These investigations indicate that GST P1-1 acts to sequester NO as DNICs, reducing their transport out of the cell by MRP1. Cell proliferation studies demonstrated the importance of the combined effect of GST P1-1 and MRP1 in protecting cells from the cytotoxic effects of NO. Thus, the DNIC storage function of GST P1-1 and ability of MRP1 to efflux DNICs are vital in protection against NO cytotoxicity. %Z FOR Codes: 30406 60104 %0 Journal Article %~ PubMed %A Suryo Rahmanto, Yohan %A Kalinowski, Danuta S %A Lane, Darius Jr %A Lok, Hiu Chuen %A Richardson, Vera %A Richardson, Des R %T Nitrogen Monoxide (NO) Storage and Transport by Dinitrosyl-Dithiol-Iron Complexes: Long-Lived NO that is Trafficked by Interacting Proteins. %B The Journal of Biological Chemistry %D 2012 %C United States %I American Society for Biochemistry and Molecular Bi %V 287 %N 10 %P 6960-6968 %@ 1083-351X %X %Z FOR Codes: 60104 60111 %0 Journal Article %~ PubMed %A Lovejoy, David B %A Sharp, Danae M %A Seebacher, Nicole %A Obeidy, Peyman %A Prichard, Thomas %A Stefani, Christian %A Basha, Maram T %A Sharpe, Philip C %A Jansson, Patric J %A Kalinowski, Danuta S %A Bernhardt, Paul V %A Richardson, Des R %T Novel Second-Generation Di-2-Pyridylketone Thiosemicarbazones Show Synergism with Standard Chemotherapeutics and Demonstrate Potent Activity against Lung Cancer Xenografts after Oral and Intravenous Administration in Vivo. %B Journal of Medicinal Chemistry %D 2012 %C United States %I American Chemical Society %V 55 %N 16 %P 7230-7244 %@ 0022-2623 %X We developed a series of second-generation di-2-pyridyl ketone thiosemicarbazone (DpT) and 2-benzoylpyridine thiosemicarbazone (BpT) ligands to improve the efficacy and safety profile of these potential antitumor agents. Two novel DpT analogues, Dp4e4mT and DpC, exhibited pronounced and selective activity against human lung cancer xenografts in vivo via the intravenous and oral routes. Importantly, these analogues did not induce the cardiotoxicity observed at high nonoptimal doses of the first-generation DpT analogue, Dp44mT. The Cu(II) complexes of these ligands exhibited potent antiproliferative activity having redox potentials in a range accessible to biological reductants. The activity of the copper complexes of Dp4e4mT and DpC against lung cancer cells was synergistic in combination with gemcitabine or cisplatin. It was demonstrated by EPR spectroscopy that dimeric copper compounds of the type [CuLCl](2), identified crystallographically, dissociate in solution to give monomeric 1:1 Cu:ligand complexes. These monomers represent the biologically active form of the complex. %Z FOR Codes: 111205 %0 Journal Article %~ PubMed %A Correnti, Colin %A Richardson, Vera %A Sia, Allyson K %A Bandaranayake, Ashok D %A Ruiz, Mario %A Suryo Rahmanto, Yohan %A KovačeviĿ, Žaklina %A Clifton, Matthew C %A Holmes, Margaret A %A Kaiser, Brett K %A Barasch, Jonathan %A Raymond, Kenneth N %A Richardson, Des R %A Strong, Roland K %T Siderocalin/Lcn2/NGAL/24p3 does not drive apoptosis through gentisic acid mediated iron withdrawal in hematopoietic cell lines. %B PLoS One %D 2012 %C United States %I Public Library of Science %V 7 %N 8 %P e43696 %@ 1932-6203 %X %Z FOR Codes: 110199 %0 Journal Article %~ PubMed %A Li, Cheng %A L??nn, Maria E %A Xu, Xiangcong %A Maghzal, Ghassan J %A Frazer, David M %A Thomas, Shane R %A Halliwell, Barry %A Richardson, Des R %A Anderson, Gregory J %A Stocker, Roland %T Sustained expression of heme oxygenase-1 alters iron homeostasis in nonerythroid cells. %B Free Radical Biology & Medicine %D 2012 %C United States %I Elsevier Inc. %V 53 %N 2 %P 366-374 %@ 0891-5849 %X Heme oxygenases initiate the catabolism of heme, releasing carbon monoxide, iron, and biliverdin. Sustained induction of heme oxygenase-1 (HO-1) in nonerythroid cells plays a key role in many pathological processes, yet the effect of long-term HO-1 expression on cellular iron metabolism in the absence of exogenous heme is poorly understood. Here we report that in a model nonerythroid cell, both transient and stable HO-1 expression increased heme oxygenase activity, but total cellular heme content was decreased only with transient enzyme expression. Sustained HO-1 activity increased the expression of both the mitochondrial iron importer mitoferrin-2 and the rate-limiting enzyme in heme synthesis, aminolevulinate synthase-1, and it augmented the mitochondrial content of heme. Also, the expression of transferrin receptor-1 and the activities of iron-regulatory proteins 1 and 2 decreased, whereas total labile iron and the regulatory activity of the heme-binding transcription factor Bach1 were unaltered. In addition, stable, but not transient, HO-1 expression decreased the activities of aconitase, as well as increasing proteasomal degradation of ferritin. Together, our results reveal a novel and coordinated adaptive response of nonerythroid cells to sustained HO-1 induction that has an impact on cellular iron homeostasis. %Z FOR Codes: 60104 %0 Journal Article %~ PubMed %A Serda, Maciej %A Kalinowski, Danuta S %A Mrozek-Wilczkiewicz, Anna %A Musiol, Robert %A Szurko, Agnieszka %A Ratuszna, Alicja %A Pantarat, Namfon %A Kovacevic, Zaklina %A Merlot, Angelica M %A Richardson, Des R %A Polanski, Jaroslaw %T Synthesis and characterization of quinoline-based thiosemicarbazones and correlation of cellular iron-binding efficacy to anti-tumor efficacy. %B Bioorganic & Medicinal Chemistry Letters %D 2012 %C United Kingdom %I Pergamon %V 22 %N 17 %P 5527-5531 %@ 1464-3405 %X %Z FOR Codes: 111204 %0 Journal Article %A Kovacevic, Zaklina %A Richardson, Desi %T Targeting Iron in Cancer Cells: A New Strategy to Inhibit Metastatic Progression %B Vitamins & Trace Elements %D 2012 %C United States %I OMICS Publishing Group %V 1 %N 3 %P 1000e114 %@ 2167-0390 %X %Z FOR Codes: 111204 %0 Journal Article %~ PubMed %A Chen, Zhiqiang %A Zhang, Daohai %A Yue, Fei %A Zheng, Minhua %A Kovacevic, Zaklina %A Richardson, Des R %T The iron chelators Dp44mT and DFO inhibit TGF-?-induced epithelial-mesenchymal transition via up-regulation of N-Myc downstream-regulated gene 1 (NDRG1). %B The Journal of Biological Chemistry %D 2012 %C United States %I American Society for Biochemistry and Molecular Bi %V 287 %N 21 %P 17016-17028 %@ 1083-351X %X The epithelial-mesenchymal transition (EMT) is a key step for cancer cell migration, invasion, and metastasis. Transforming growth factor-? (TGF-?) regulates the EMT and the metastasis suppressor gene, N-myc downstream-regulated gene-1 (NDRG1), could play a role in regulating the TGF-? pathway. NDRG1 expression is markedly increased after chelator-mediated iron depletion via hypoxia-inducible factor 1?-dependent and independent pathways (Le, N. T. and Richardson, D. R. (2004) Blood 104, 2967-2975). Moreover, novel iron chelators show marked and selective anti-tumor activity and are a potential new class of anti-metabolites. Considering this, the current study investigated the relationship between NDRG1 and the EMT to examine if iron chelators can inhibit the EMT via NDRG1 up-regulation. We demonstrated that TGF-? induces the EMT in HT29 and DU145 cells. Further, the chelators, desferrioxamine (DFO) and di-2-pyridylketone-4,4-dimethyl-3-thiosemicarbazone (Dp44mT), inhibited the TGF-?-induced EMT by maintaining E-cadherin and ?-catenin, at the cell membrane. We then established stable clones with NDRG1 overexpression and knock-down in HT29 and DU145 cells. These data showed that NDRG1 overexpression maintained membrane E-cadherin and ?-catenin and inhibited TGF-?-stimulated cell migration and invasion. Conversely, NDRG1 knock-down caused morphological changes from an epithelial- to fibroblastic-like phenotype and also increased migration and invasion, demonstrating NDRG1 knockdown induced the EMT and enhanced TGF-? effects. We also investigated the mechanisms involved and showed the TGF-?/SMAD and Wnt pathways were implicated in NDRG1 regulation of E-cadherin and ?-catenin expression and translocation. This study demonstrates that chelators inhibit the TGF-?-induced EMT via a process consistent with NDRG1 up-regulation and elucidates the mechanism of their activity. %Z FOR Codes: 111201 111201 %0 Journal Article %~ PubMed %A Lovejoy, David B %A Jansson, Patric J %A Brunk, Ulf T %A Wong, Jacky %A Ponka, Prem %A Richardson, Des R %T Antitumor Activity of Metal-Chelating Compound Dp44mT Is Mediated by Formation of a Redox-Active Copper Complex That Accumulates in Lysosomes %B Cancer research %D 2011 %C United States %I American Association for Cancer Research (A AC R) %V 71 %N 17 %P 5871-80 %@ 0008-5472 %X The metal-chelating compound Dp44mT is a di-2-pyridylketone thiosemicarbazone (DpT) which displays potent and selective antitumor activity. This compound is receiving translational attention, but its mechanism is poorly understood. Here, we report that Dp44mT targets lysosome integrity through copper binding. Studies using the lysosomotropic fluorochrome acridine orange established that the copper-Dp44mT complex (Cu[Dp44mT]) disrupted lysosomes. This targeting was confirmed with pepstatin A-BODIPY FL, which showed redistribution of cathepsin D to the cytosol with ensuing cleavage of the proapoptotic BH3 protein Bid. Redox activity of Cu[Dp44mT] caused cellular depletion of glutathione, and lysosomal damage was prevented by cotreatment with the glutathione precursor N-acetylcysteine. Copper binding was essential for the potent antitumor activity of Dp44mT, as coincubation with nontoxic copper chelators markedly attenuated its cytotoxicity. Taken together, our studies show how the lysosomal apoptotic pathway can be selectively activated in cancer cells by sequestration of redox-active copper. Our findings define a novel generalized strategy to selectively target lysosome function for chemotherapeutic intervention against cancer. %Z FOR Codes: 111501 111205 %0 Journal Article %~ PubMed %A Yu, Yu %A Richardson, Des R %T Cellular Iron Depletion Stimulates the JNK and p38 MAPK Signaling Transduction Pathways, Dissociation of ASK1-Thioredoxin, and Activation of ASK1. %B Journal of Biological Chemistry %D 2011 %C United States %I American Society for Biochemistry and Molecular Bi %V 286 %N 17 %P 15413-15427 %@ 1083-351X %X The role of signaling pathways in the regulation of cellular iron metabolism is becoming increasingly recognized. Iron chelation is used for the treatment of iron overload but also as a potential strategy for cancer therapy, because iron depletion results in cell cycle arrest and apoptosis. This study examined potential signaling pathways affected by iron depletion induced by desferrioxamine (DFO) or di-2-pyridylketone-4,4-dimethyl-3-thiosemicarbazone (Dp44mT). Both chelators affected multiple molecules in the mitogen-activated protein kinase (MAPK) pathway, including a number of dual specificity phosphatases that directly de-phosphorylate MAPKs. Examination of the phosphorylation of major MAPKs revealed that DFO and Dp44mT markedly increased phosphorylation of stress-activated protein kinases, JNK and p38, without significantly affecting the extracellular signal-regulated kinase (ERK). Redox-inactive DFO-iron complexes did not affect phosphorylation of JNK or p38, whereas the redox-active Dp44mT-iron complex significantly increased the phosphorylation of these kinases similarly to Dp44mT alone. Iron or N-acetylcysteine supplementation reversed Dp44mT-induced up-regulation of phospho-JNK, but only iron was able to reverse the effect of DFO on JNK. Both iron chelators significantly reduced ASK1-thioredoxin complex formation, resulting in the increased phosphorylation of ASK1, which activates the JNK and p38 pathways. Thus, dissociation of ASK1 could serve as an important signal for the phosphorylation of JNK and p38 activation observed after iron chelation. Phosphorylation of JNK and p38 likely play an important role in mediating the cell cycle arrest and apoptosis induced by iron depletion. %Z FOR Codes: 60111 %0 Journal Article %~ PubMed %A Saletta, Federica %A Suryo Rahmanto, Yohan %A Siafakas, Aritee R %A Richardson, Des R %T Cellular iron-depletion and the mechanisms involved in the iron-dependent regulation of the growth arrest and DNA damage family of genes. %B The Journal of biological chemistry %D 2011 %C United States %I American Society for Biochemistry and Molecular Bi %V 286 %N 41 %P 35396-406 %@ 1083-351X %X Iron plays a crucial part in proliferation while iron deficiency results in G(1)/S arrest, DNA damage, and apoptosis. However, the precise role of iron in cell cycle control remains unclear. We showed that iron depletion using the iron chelators, desferrioxamine (DFO), or 2-hydroxy-1-napthylaldehyde isonicotinoyl hydrazone (311), increased the mRNA levels of the growth arrest and DNA damage 45?? gene, GADD45?? (Darnell, G. and Richardson, D. R. (1999) Blood 94, 781-792). In this study, we examined the effect of iron depletion on up-regulating GADD family members involved in growth control, including cell cycle arrest, apoptosis, and DNA repair, making them therapeutic targets for tumor suppression. We showed the GADD family members were up-regulated by cellular iron depletion. Further, up-regulation of GADD45?? after iron deprivation was independent of hypoxia-inducible factor-1?? (HIF-1??), octamer-1 (Oct-1), p53 and early growth response 1 (Egr1). We then analyzed the regulatory elements responsible for iron depletion-mediated regulation of GADD45?? and identified the specific transcription factor/s involved. This region was within -117 bp and -81 bp relative to the start codon where the consensus sequences of three transcription factors are located: the CCAAT-binding factor/nuclear factor-Y (NF-Y), the stabilizing molecule v-MYB and the enhancer, CCAAT enhancer-binding protein (CEBP??). Mutation analysis, shRNA studies, Western blotting, and electrophoretic mobility shift assays led to the identification of NF-Y in the transcriptional up-regulation of GADD45?? after iron depletion. Furthermore, like GADD45??, NF-YA was up-regulated after iron chelation and down-regulated by iron supplementation. These results are important for understanding the mechanisms of iron depletion-mediated cell cycle arrest, DNA damage repair, and apoptosis. %Z FOR Codes: 60199 110199 %0 Journal Article %~ PubMed %A Kovacevic, Zaklina %A Yu, Yu %A Richardson, Des R %T Chelators to the rescue: different horses for different courses! %B Chemical Research in Toxicology %D 2011 %C United States %I American Chemical Society %V 24 %N 3 %P 279-282 %@ 1520-5010 %X %Z FOR Codes: 30401 111204 111502 %0 Journal Article %~ PubMed %A Zhang, Daohai %A Richardson, Des R %T Endoplasmic Reticulum Protein 29 (ERp29): The emerging role in cancer. %B The international journal of biochemistry & cell biology %D 2011 %C United Kingdom %I Pergamon %V 43 %N 1 %P 33-6 %@ 1357-2725 %X The endoplasmic reticulum protein 29 (ERp29) is a molecule that facilitates processing and transport of proteins in the early secretory pathway. Structural and functional analyses have suggested a biological role as a putative chaperone in the endoplasmic reticulum. The N-terminal domain of ERp29 resembles the thioredoxin domain of protein disulfide isomerase, but lacks its redox-active function due to the absence of an active motif consisting of double cysteines. In the context of carcinogenesis, the role of ERp29 in cancer progression has not been fully elucidated. However, recent studies indicate that high expression of ERp29 inversely correlates to tumor progression. In addition, over-expression of ERp29 significantly inhibits proliferation and suppresses tumorigenesis by modulating ER stress signaling and the mesenchymal-epithelial transition in breast cancer cells. In this review, we summarize the biological properties of ERp29 and its novel function as a tumor suppressor. %Z FOR Codes: 111201 %0 Journal Article %~ PubMed %A Stefani, Christian %A Punnia-Moorthy, Gaya %A Lovejoy, David B %A Jansson, Patric J %A Kalinowski, Danuta S %A Sharpe, Philip C %A Bernhardt, Paul V %A Richardson, Des R %T Halogenated 2'-Benzoylpyridine Thiosemicarbazone (XBpT) Chelators with Potent and Selective Anti-Neoplastic Activity: Relationship to Intracellular Redox Activity. %B Journal of medicinal chemistry %D 2011 %C United States %I American Chemical Society %V 54 %N 19 %P 6936-48 %@ 0022-2623 %X Iron chelators of the 2''-benzoylpyridine thiosemicarbazone (BpT) class show substantial potential as anticancer agents. To explore structure-activity relationships, new BpT analogues were designed that incorporated halogen substituents on the noncoordinating phenyl group (XBpTs). These XBpT ligands exhibited potent antiproliferative activity with some analogues exceeding that of the parent BpT compound. Importantly, there was an appreciable therapeutic index in vitro, as mortal cells were significantly less affected by these chelators relative to neoplastic cells. The addition of a halogen led to a halogen-specific increase in the redox potential of XBpT-Fe complexes. Probing for chelator-induced intracellular reactive oxygen species (ROS) with the fluorescent probe, 2'',7''-dichlorofluorescein, revealed a 1.5-4.7-fold increase in fluorescence upon incorporation of Cl, Br, or I to the parent analogues. Furthermore, an important structure-activity relationship was deduced where the addition of halogens led to a positive correlation between intracellular ROS generation and antiproliferative activity in the more hydrophilic BpT parent compounds. %Z FOR Codes: 30401 %0 Journal Article %~ PubMed %A Debebe, Zufan %A Ammosova, Tatyana %A Breuer, Denitra %A Lovejoy, David B %A Kalinowski, Danuta S %A Karla, Pradeep K %A Kumar, Krishna %A Jerebtsova, Marina %A Ray, Patricio %A Kashanchi, Fatah %A Gordeuk, Victor R %A Richardson, Des R %A Nekhai, Sergei %T Iron chelators of the di-2-pyridylketone thiosemicarbazone and 2-benzoylpyridine thiosemicarbazone series inhibit HIV-1 transcription: identification of novel cellular targets--iron, cyclin-dependent kinase (CDK) 2, and CDK9. %B Molecular Pharmacology %D 2011 %C United States %I American Society for Pharmacology and Experimental Therapeutics %V 79 %N 1 %P 185-196 %@ 1521-0111 %X HIV-1 transcription is activated by HIV-1 Tat protein, which recruits cyclin-dependent kinase 9 (CDK9)/cyclin T1 and other host transcriptional coactivators to the HIV-1 promoter. Tat itself is phosphorylated by CDK2, and inhibition of CDK2 by small interfering RNA, the iron chelator 2-hydroxy-1-naphthylaldehyde isonicotinoyl hydrazone (311), and the iron chelator deferasirox (ICL670) inhibits HIV-1 transcription. Here we have analyzed a group of novel di-2-pyridylketone thiosemicarbazone- and 2-benzoylpyridine thiosemicarbazone-based iron chelators that exhibit marked anticancer activity in vitro and in vivo (Proc Natl Acad Sci USA 103:7670-7675, 2006; J Med Chem 50:3716-3729, 2007). Several of these iron chelators, in particular 2-benzoylpyridine 4-allyl-3-thiosemicarbazone (Bp4aT) and 2-benzoylpyridine 4-ethyl-3-thiosemicarbazone (Bp4eT), inhibited HIV-1 transcription and replication at much lower concentrations than did 311 and ICL670. Neither Bp4aT nor Bp4eT were toxic after a 24-h incubation. However, longer incubations for 48 h or 72 h resulted in cytotoxicity. Analysis of the molecular mechanism of HIV-1 inhibition showed that the novel iron chelators inhibited basal HIV-1 transcription, but not the nuclear factor-?B-dependent transcription or transcription from an HIV-1 promoter with inactivated SP1 sites. The chelators inhibited the activities of CDK2 and CDK9/cyclin T1, suggesting that inhibition of CDK9 may contribute to the inhibition of HIV-1 transcription. Our study suggests the potential usefulness of Bp4aT or Bp4eT in antiretroviral regimens, particularly where resistance to standard treatment occurs. %Z FOR Codes: 111504 30401 %0 Journal Article %~ PubMed %A Bernhardt, Paul V %A Richardson, Des R %T Metal chelation. %B Current Topics in Medicinal Chemistry %D 2011 %C Netherlands %I Bentham Science Publishers Ltd. %V 11 %N 5 %P 482 %@ 1873-4294 %X %Z FOR Codes: 60104 %0 Journal Article %~ PubMed %A Huang, Michael Li-Hsuan %A Lane, Darius J R %A Richardson, Des R %T Mitochondrial Mayhem: The Mitochondrion as a Modulator of Iron Metabolism and Its Role in Disease. %B Antioxidants & redox signaling %D 2011 %C United States %I Mary Ann Liebert, Inc. Publishers %V 15 %N 12 %P 3003-19 %@ 1557-7716 %X The mitochondrion plays vital roles in various aspects of cellular metabolism, ranging from energy transduction and apoptosis to the synthesis of important molecules such as heme. Mitochondria are also centrally involved in iron metabolism, as exemplified by disruptions in mitochondrial proteins that lead to perturbations in whole-cell iron processing. Recent investigations have identified a host of mitochondrial proteins (e.g., mitochondrial ferritin; mitoferrins 1 and 2; ABCBs 6, 7, and 10; and frataxin) that may play roles in the homeostasis of mitochondrial iron. These mitochondrial proteins appear to participate in one or more processes of iron storage, iron uptake, and heme and iron-sulfur cluster synthesis. In this review, we present and critically discuss the evidence suggesting that the mitochondrion may contribute to the regulation of whole-cell iron metabolism. Further, human diseases that arise from a dysregulation of these mitochondrial molecules reveal the ability of the mitochondrion to communicate with cytosolic iron metabolism to coordinate whole-cell iron processing and to fulfill the high demands of this organelle for iron. This review highlights new advances in understanding iron metabolism in terms of novel molecular players and diseases associated with its dysregulation. %Z FOR Codes: 60104 %0 Journal Article %~ PubMed %A Kovacevic, Zaklina %A Chikhani, Sherin %A Lovejoy, David B %A Richardson, Des R %T Novel thiosemicarbazone iron chelators induce up-regulation and phosphorylation of the metastasis suppressor, NDRG1: A new strategy for the treatment of pancreatic cancer %B Molecular pharmacology %D 2011 %C United States %I American Society for Pharmacology and Experimental Therapeutics %V 80 %N 4 %P 598-609 %@ 1521-0111 %X Pancreatic cancer is an aggressive neoplasm, with a mortality rate close to 100%. The most successful agent for pancreatic cancer treatment is gemcitabine, although the overall effect in terms of patient survival remains very poor. This study was initiated to evaluate a novel class of anticancer agents against pancreatic cancer. This group of compounds belongs to the dipyridyl thiosemicarbazone class that have been shown to have potent and selective activity against a range of different neoplasms in vitro and in vivo. We demonstrate for the first time in pancreatic cancer that these agents increase the expression of the growth and metastasis suppressor N-myc downstream-regulated gene 1 and its phosphorylation at Ser330 and Thr346 that is important for its activity against this tumor. In addition, these agents increased expression of the cyclin-dependent kinase inhibitor p21(CIP1/WAF1), whereas decreasing cyclin D1 in pancreatic cancer cells. Together, these molecular alterations account, in part, for the pronounced antitumor activity observed. Indeed, these agents had significantly higher antiproliferative activity in vitro than the established treatments for pancreatic cancer, namely gemcitabine and 5-fluorouracil. Studies in vivo demonstrated that a novel thiosemicarbazone, namely di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone hydrochloride, completely inhibited the growth of pancreatic cancer xenografts with no evidence of marked alterations in normal tissue histology. Together, our studies have identified molecular effectors of a novel and potent antitumor agent that could be useful for pancreatic cancer treatment. %Z FOR Codes: 111501 %0 Journal Article %~ PubMed %A Saletta, Federica %A Kovacevic, Zaklina %A Richardson, Des Raymond %T Research Spotlight: Iron chelation: deciphering novel molecular targets for cancer therapy. The tip of the iceberg of a web of iron-regulated molecules. %B Future Medicinal Chemistry %D 2011 %C United Kingdom %I Future Science Ltd. %V 3 %N 16 %P 1983-1986 %@ 1756-8927 %X The response of cells to cellular iron depletion is complex with multiple molecules and signaling pathways being involved. Indeed, this is far broader than just the effect on the classical target, ribonucleotide reductase. It is likely that a network of interactions exist between the molecular players and that the relationships currently known only represent the ''tip of an iceberg'' in terms of understanding the response of cells to iron deprivation. This article describes some of the research being undertaken in this area by the Richardson group at the Univerity of Syndney, Australia. %Z FOR Codes: 30499 111501 %0 Journal Article %~ PubMed %A Sharpe, Philip C %A Richardson, Des R %A Kalinowski, Danuta S %A Bernhardt, Paul V %T Synthetic and natural products as iron chelators. %B Current Topics in Medicinal Chemistry %D 2011 %C Netherlands %I Bentham Science Publishers Ltd. %V 11 %N 5 %P 591-607 %@ 1873-4294 %X An evaluation of existing and proposed Fe chelators, both synthetic and natural products, for the treatment of Fe-overload disease must address a number of issues. There are fundamental parameters that determine the efficacy of a drug: absorption, distribution, metabolism, clearance and toxicity. However, the administration of chelator for Fe overload aims to generate Fe complexes in vivo that are able to be excreted. Hence, the chemical and pharmacological properties of the complexes formed are equally important as the chelators themselves. The redox properties of the Fe complexes formed is particularly relevant to their toxicity. If both Fe(II) and Fe(III) oxidation states of the complexes are biologically accessible, then there is potential for the auto-catalytic production of deleterious free radicals, by Fenton-type chemistry. In addition, since the burden of Fe overload disease falls predominantly on some of the poorest economies, the cost of a drug must be considered, as well as the mode of delivery. There are also possible issues with the use of naturally occurring ligands, which may form Fe complexes capable of being utilised by opportunistic bacteria. This review will concentrate on recent developments in our chemical understanding of existing chelators approved or proposed for use and will also consider some of the candidates from natural sources that have been recently proposed. %Z FOR Codes: 111599 %0 Journal Article %~ PubMed %A Kovacevic, Zaklina %A Kalinowski, Danuta S %A Lovejoy, David B %A Yu, Yu %A Rahmanto, Yohan Suryo %A Sharpe, Phillip C %A Bernhardt, Paul V %A Richardson, Des R %T The Medicinal Chemistry of Novel Iron Chelators for the Treatment of Cancer. %B Current topics in medicinal chemistry %D 2011 %C Netherlands %I Bentham Science Publishers Ltd. %V 11 %N 5 %P 483-99 %@ 1873-4294 %X Cancer is one of the leading causes of death worldwide and there is an increasing need for novel anti-tumor therapeutics with greater selectivity and potency. A new strategy in the treatment of cancer has focused on targeting an essential cell metabolite, iron (Fe). Iron is vital for cell growth and metabolism, forming a crucial component of the active site of ribonucleotide reductase (RR), the rate-limiting enzyme in DNA synthesis. Cancer cells in particular require large amounts of Fe to proliferate, making them more susceptible to the Fe deficiency caused by Fe chelators. Beginning with primordial siderophores, Fe chelators have since evolved to a new generation of potent and efficient anti-cancer agents. Recently, investigations have led to the generation of novel di-2-pyridylketone thiosemicarbazone (DpT) and 2-benzoylpyridine thiosemicarbazone (BpT) ligands that demonstrate marked and selective anti-tumor activity both in vitro and in vivo against a wide spectrum of tumors. The mechanism of action of these novel ligands includes alterations in the expression of key regulatory molecules as well as the generation of redox active Fe complexes. Interestingly, non-synthetic Fe chelators including silybin and curcumin, both of which are derived from plants, also have vast potential in the treatment of cancer. This review explores the development of novel Fe chelators for the treatment of cancer and their mechanisms of action. %Z FOR Codes: 30499 111204 %0 Journal Article %~ PubMed %A Yu, Yu %A Suryo Rahmanto, Yohan %A Hawkins, Clare L %A Richardson, Des R %T The Potent and Novel Thiosemicarbazone Chelators, Dp44mT and Bp44mT, Affect Crucial Thiol Systems Required for Ribonucleotide Reductase Activity. %B Molecular pharmacology %D 2011 %C United States %I American Society for Pharmacology and Experimental Therapeutics %V 79 %N 6 %P 921-31 %@ 1521-0111 %X Di-2-pyridylketone-4,4-dimethyl-3-thiosemicarbazone possesses potent and selective antitumor activity. Its cytotoxicity has been attributed to iron chelation leading to inhibition of the iron-containing enzyme ribonucleotide reductase (RR). Thiosemicarbazone iron complexes have been shown to be redox-active, although their effect on cellular antioxidant systems is unclear. Using a variety of antioxidants, we found that only N-acetylcysteine significantly inhibited thiosemicarbazone-induced antiproliferative activity. Thus, we examined the effects of thiosemicarbazones on major thiol-containing systems considering their key involvement in providing reducing equivalents for RR. Thiosemicarbazones significantly (p < 0.001) elevated oxidized trimeric thioredoxin levels to 213 ?? 5% (n = 3) of the control. This was most likely due to a significant (p < 0.01) decrease in thioredoxin reductase activity to 65 ?? 6% (n = 4) of the control. We were surprised to find that the non-redox-active chelator desferrioxamine increased thioredoxin oxidation to a lower extent (152 ?? 9%; n = 3) and inhibited thioredoxin reductase activity (62 ?? 5%; n = 4), but at a 10-fold higher concentration than thiosemicarbazones. In contrast, only the thiosemicarbazones significantly (p < 0.05) reduced the glutathione/oxidized-glutathione ratio and the activity of glutaredoxin that requires glutathione as a reductant. All chelators significantly decreased RR activity, whereas the NADPH/NADP(total) ratio was not reduced. This was important to consider because NADPH is required for thiol reduction. Thus, thiosemicarbazones could have an additional mechanism of RR inhibition via their effects on major thiol-containing systems. %Z FOR Codes: 111501 30401 %0 Journal Article %~ PubMed %A Kovacevic, Zaklina %A Sivagurunathan, Sutharshani %A Mangs, Helena %A Chikhani, Sherin %A Zhang, Daohai %A Richardson, Des R %T The metastasis suppressor, N-MYC downstream regulated gene 1 (NDRG1), UP-regulates P21 VIA P53-independent mechanisms. %B Carcinogenesis %D 2011 %C United Kingdom %I Oxford University Press %V 32 %N 5 %P 732-40 %@ 1460-2180 %X The metastasis suppressor, N-myc downstream regulated gene-1 (NDRG1), has been shown to markedly reduce metastasis of numerous tumors. The current study was focused on further elucidating the molecular mechanisms behind the antitumor function of NDRG1. We have identified for the first time that NDRG1 upregulates the potent cyclin-dependent kinase inhibitor, p21. This effect was observed in three different cancer cell types, including PC3MM and DU145 prostate cancer cells and H1299 lung carcinoma cells, and occurred independently of p53. In addition, reducing NDRG1 expression using short hairpin RNA in PC3MM and DU145 cells resulted in significantly reduced p21 protein levels. Hence, p21 is closely correlated with NDRG1 expression in these latter cell types. Examining the mechanisms behind the effect of NDRG1 on p21 expression, we found that NDRG1 upregulated p21 via transcriptional and posttranscriptional mechanisms in prostate cancer cells, although its effect on H1299 cells was posttranscriptional only. Further studies identified two additional NDRG1 protein targets. The dominant-negative p63 isoform, ??Np63, which has been found to inhibit p21 transcription, was downregulated by NDRG1. On the other hand, a truncated 50 kDa MDM2 isoform (p50(MDM2)), which may protect p21 from proteasomal degradation, was upregulated by NDRG1. The downregulation of ??Np63 and upregulation of p50(MDM2) are potential mechanisms by which NDRG1 increases p21 expression in these cells. Additional functional studies identified that NDRG1 inhibits cancer cell migration, suggesting that p21 is a molecular player in its antimetastatic activity. %Z FOR Codes: 111207 60104 %0 Journal Article %~ Isi %A Miao, Q. H. %A Xu, D. X. %A Wang, Z. %A Xu, L. %A Wang, T. W. %A Wu, Y. %A Lovejoy, D. B. %A Kalinowski, D. S. %A Richardson, D. R. %A Nie, G. J. %A Zhao, Y. L. %T Amphiphilic hyper-branched co-polymer nanoparticles for the controlled delivery of anti-tumor agents %B Biomaterials %D 2010 %C Netherlands %I Elsevier BV %V 31 %N 28 %P 7364-7375 %@ 0142-9612 %X %Z FOR Codes: 111205 111501 %0 Book Section %A Kalinowski, Danuta %A Richardson, Des %T Cellular and Molecular Biology of Iron-Binding Proteins %B Cellular and Molecular Biology of Metals %D 2010 %C United States %I Taylor and Francis %V %N %P 167-180 %@ 9781420059977 %E Zalups, Rudolfs K. %E Koropatnick, D. James %X %Z FOR Codes: 60109 %0 Journal Article %~ PubMed %A Bendova, Petra %A Mackova, Eliska %A Haskova, Pavlina %A Vavrova, Anna %A Jirkovsky, Eduard %A Sterba, Martin %A Popelova, Olga %A Kalinowski, Danuta S %A Kovarikova, Petra %A Vavrova, Katerina %A Richardson, Des R %A Simunek, Tomas %T Comparison of Clinically Used and Experimental Iron Chelators for Protection against Oxidative Stress-Induced Cellular Injury. %B Chemical research in toxicology %D 2010 %C United States %I American Chemical Society %V 23 %N 6 %P 1105-14 %@ 1520-5010 %X Iron imbalance plays an important role in oxidative stress associated with numerous pathological conditions. Therefore, iron chelation may be an effective therapeutic approach, but progress in this area is hindered by the lack of effective ligands. Also, the potential favorable effects of chelators against oxidative injury have to be balanced against their own toxicity due to iron depletion and the ability to generate redox-active iron complexes. In this study, we compared selected iron chelators (both drugs used in clinical practice as well as experimental agents) for their efficacy to protect cells against model oxidative injury induced by tert-butyl hydroperoxide (t-BHP). In addition, intracellular chelation efficiency, redox activity, and the cytotoxicity of the chelators and their iron complexes were assayed. Ethylenediaminetetraacetic acid failed to protect cells against t-BHP cytotoxicity, apparently due to the redox activity of the formed iron complex. Hydrophilic desferrioxamine exerted some protection but only at very high clinically unachievable concentrations. The smaller and more lipophilic chelators, deferiprone, deferasirox, and pyridoxal isonicotinoyl hydrazone, were markedly more effective at preventing oxidative injury of cells. The most effective chelator in terms of access to the intracellular labile iron pool was di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone. However, overall, the most favorable properties in terms of protective efficiency against t-BHP and the chelator''s own inherent cytotoxicity were observed with salicylaldehyde isonicotinoyl hydrazone. This probably relates to the optimal lipophilicity of this latter agent and its ability to generate iron complexes that do not induce marked redox activity. %Z FOR Codes: 304 %0 Journal Article %~ PubMed %A Liu, Joe %A Obando, Daniel %A Schipanski, Liam G %A Groebler, Ludwig K %A Witting, Paul K %A Kalinowski, Danuta S %A Richardson, Des R %A Codd, Rachel %T Conjugates of Desferrioxamine B (DFOB) with Derivatives of Adamantane or with Orally Available Chelators as Potential Agents for Treating Iron Overload. %B Journal of medicinal chemistry %D 2010 %C United States %I American Chemical Society %V 53 %N 3 %P 1370-82 %@ 0022-2623 %X Desferrioxamine B (DFOB) conjugates with adamantane-1-carboxylic acid, 3-hydroxyadamantane-1-carboxylic acid, 3,5-dimethyladamantane-1-carboxylic acid, adamantane-1-acetic acid, 4-methylphenoxyacetic acid, 3-hydroxy-2-methyl-4-oxo-1-pyridineacetic acid (N-acetic acid derivative of deferiprone), or 4-[3,5-bis(2-hydroxyphenyl)-1,2,4-triazol-1-yl]benzoic acid (deferasirox) were prepared and the integrity of Fe(III) binding of the compounds was established from electrospray ionization mass spectrometry and RP-HPLC measurements. The extent of intracellular (59)Fe mobilized by the DFOB-3,5-dimethyladamantane-1-carboxylic acid adduct was 3-fold greater than DFOB alone, and the IC(50) value of this adduct was 6- or 15-fold greater than DFOB in two different cell types. The relationship between logP and (59)Fe mobilization for the DFOB conjugates showed that maximal mobilization of intracellular (59)Fe occurred at a logP value approximately 2.3. This parameter, rather than the affinity for Fe(III), appears to influence the extent of intracellular (59)Fe mobilization. The low toxicity-high Fe mobilization efficacy of selected adamantane-based DFOB conjugates underscores the potential of these compounds to treat iron overload disease in patients with transfusional-dependent disorders such as beta-thalassemia. %Z FOR Codes: 30405 30404 110103 %0 Journal Article %~ PubMed %A Stariat, Jn %A Kovarkov, Petra %A Klimes, Jir %A Kalinowski, Danuta S %A Richardson, Des R %T Development of an LC-MS/MS method for analysis of interconvertible Z/E isomers of the novel anticancer agent, Bp4eT. %B Analytical and Bioanalytical Chemistry %D 2010 %C Germany %I Springer %V 397 %N 1 %P 161-171 %@ 1618-2650 %X This study was focused on a liquid chromatography/tandem mass spectrometry (LC/MS/MS) method development for quantification of a novel potential anticancer agent, 2-benzoylpyridine 4-ethyl-3-thiosemicarbazone (Bp4eT), in aqueous media. Solid Bp4eT was found to consist predominantly of the Z isomer, while in aqueous media, both isomers coexist. Sufficient separation of both isomers was achieved on a Synergi 4u Polar RP column with a mobile phase composed of 2 mM ammonium formate, acetonitrile, and methanol (30:63:7; v/v/v). The photo diode array analysis of both isomers demonstrated different absorption spectra which hindered UV-based quantification. However, an equal and reproducible response was found for both isomers using an MS detector, which enables the determination of the total content of Bp4eT (i.e., both E- and Z- isomeric forms) by summation of the peak areas of both isomers. 2-Hydroxy-1-naphthylaldehyde 4-methyl-3-thiosemicarbazone (N4mT) was selected as the internal standard. Quantification was performed in selective reaction monitoring using the main fragments of [M+H](+) (240 m/z for Bp4eT and 229 m/z for N4mT). The method was validated over 20-600 ng/ml. This procedure was applied to a preformulation study to determine the proper vehicle for parenteral administration. It was found that Bp4eT was poorly soluble in aqueous media. However, the solubility can be effectively improved using pharmaceutical cosolvents. In fact, a 1:1 mixture of PEG 300/0.14 M saline markedly increased solubility and may be a useful drug formulation for intravenous administration. This investigation further accelerates development of novel anticancer thiosemicarbazones. The described methods will be useful for analogs currently under development and suffering the same analytical issue. %Z FOR Codes: 111204 110316 110316 %0 Journal Article %~ PubMed %A Lane, Darius J R %A Richardson, Des R %T Frataxin, a molecule of mystery: trading stability for function in its iron-binding site. %B Biochemical Journal %D 2010 %C United States %I Portland Press Ltd. %V 426 %N 2 %P e1-e3 %@ 1470-8728 %X What are the structural implications for iron binding by frataxin, the mitochondrial protein whose decreased expression results in Friedreich''s ataxia? Though frataxin has been shown to be essential for proper handling of iron within mitochondria (e.g. for iron-sulfur cluster and haem biosynthesis), its exact molecular function remains unclear. In this issue of the Biochemical Journal, Correia and colleagues. investigate the relationship between structure and function at the putative iron-binding site of Yfh1 (yeast frataxin). Using a host of Yfh1 combination point mutants, the authors observe that the presence of a semi-conserved pocket of negative charge within the ''acidic ridge'' region (thought to be responsible for iron binding) only mildly enhances Yfh1''s ability to bind iron, though it does significantly increase the protein''s structural flexibility. The general emerging view is that frataxin''s keystone role in mitochondrial iron metabolism depends on iron binding. This appears to have downstream effects on protein-protein interactions that are crucial for frataxin function. The current results reveal a somewhat delicate relationship between iron binding and structural plasticity that may help unravel the enigma of frataxin''s metabolic roles. %Z FOR Codes: 60106 %0 Journal Article %~ Isi %A Otevrel, J. %A Mandelova, Z. %A Pesko, M. %A Guo, J. H. %A Kralova, K. %A Sersen, F. %A Vejsova, M. %A Kalinowski, D. S. %A Kovacevic, Z. %A Coffey, A. %A Csollei, J. %A Richardson, D. R. %A Jampilek, J. %T Investigating the Spectrum of Biological Activity of Ring-Substituted Salicylanilides and Carbamoylphenylcarbamates %B Molecules %D 2010 %C Switzerland %I Molecular Diversity Preservation International (M %V 15 %N 11 %P 8122-8142 %@ 1420-3049 %X %Z FOR Codes: 30401 %0 Journal Article %~ Isi %A Brychtova, K. %A Opatrilova, R. %A Raich, I. %A Kalinowski, D. S. %A Dvorakova, L. %A Placek, L. %A Csollei, J. %A Richardson, D. R. %A Jampilek, J. %T Investigating the activity of 2-substituted alkyl-6-(2,5-dioxopyrrolidin-1-yl)hexanoates as skin penetration enhancers %B Bioorganic & Medicinal Chemistry %D 2010 %C United Kingdom %I Pergamon %V 18 %N 24 %P 8556-8565 %@ 0968-0896 %X %Z FOR Codes: 30401 %0 Journal Article %~ PubMed %A Mrozek-Wilczkiewicz, Anna %A Kalinowski, Danuta S %A Musiol, Robert %A Finster, Jacek %A Szurko, Agnieszka %A Serafin, Katarzyna %A Knas, Magdalena %A Kamalapuram, Sishir K %A Kovacevic, Zaklina %A Jampilek, Josef %A Ratuszna, Alicja %A Rzeszowska-Wolny, Joanna %A Richardson, Des R %A Polanski, Jaroslaw %T Investigating the anti-proliferative activity of styrylazanaphthalenes and azanaphthalenediones. %B Bioorganic & medicinal chemistry %D 2010 %C United Kingdom %I Pergamon %V 18 %N 7 %P 2664-2671 %@ 0968-0896 %X A group of styrylazanaphthalenes and azanaphthalenediones were synthesized and tested for their anti-proliferative activity. Most of the compounds were obtained with the use of microwave-assisted synthesis. The lipophilicity of the compounds was measured by RP-HPLC and their anti-proliferative activity was assayed against the human SK-N-MC neuroepithelioma and HCT116 human colon carcinoma cell lines. Active compounds were also tested in clonogenity and comet assays. Several quinazolinone and styrylquinazoline analogues were found to have markedly greater anti-proliferative activity than desferoxamine and cis-platin. %Z FOR Codes: 110316 %0 Journal Article %~ PubMed %A Kovacevic, Z %A Kalinowski, D S %A Lovejoy, D B %A Quach, P %A Wong, J %A Richardson, D R %T Iron Chelators: Development of Novel Compounds with High and Selective Anti-Tumour Activity. %B Current Drug Delivery %D 2010 %C Netherlands %I Bentham Science Publishers Ltd. %V 7 %N 3 %P 194-207 %@ 1875-5704 %X Targeting essential nutrients (eg., those required for DNA synthesis) to inhibit cancer cell growth is a well established therapeutic strategy. A good example is the highly successful folate antagonist, methotrexate. However, up until recently, strategies to target iron which is also crucial for DNA synthesis have not been systematically explored to develop agents for the treatment of cancer. Over the last 15 years, our laboratory has embarked upon structure-activity studies designed to develop novel Fe chelators with anti-cancer efficacy. These studies have led to the development of the dipyridyl thiosemicarbazone chelators that show potent and selective anti-cancer activity and which overcome resistance to other cytotoxic agents. This class of compounds include the chelator, di-2-pyridylketone-4,4-dimethyl-3-thiosemicarbazone (Dp44mT), which at optimal doses markedly inhibits tumour growth and is well tolerated. Moreover, this ligand does not induce overt Fe-depletion in vivo, probably because very low doses (0.4 mg/kg) are effective at inhibiting tumour growth. Importantly, our compounds are far more active and less toxic than the chelator, Triapine(R), that is being assessed in a wide variety of international clinical trials. A vital part of the mechanism of action of these compounds is their ability to form a redox-active Fe complex that generates radicals to inhibit tumour growth. Due to their relatively high lipophilicity and low molecular weight of this class of compounds, oral activity may be expected in addition to their well known efficacy via the intravenous route. %Z FOR Codes: 30401 %0 Journal Article %~ PubMed %A Merlot, Angelica M %A Pantarat, Namfon %A Lovejoy, David B %A Kalinowski, Danuta S %A Richardson, Des R %T Membrane Transport and Intracellular Sequestration of Novel Thiosemicarbazone Chelators for the Treatment of Cancer. %B Molecular pharmacology %D 2010 %C United States %I American Society for Pharmacology and Experimental Therapeutics %V 78 %N 4 %P 675-84 %@ 1521-0111 %X Iron is a critical nutrient for DNA synthesis and cellular proliferation. Targeting iron in cancer cells using specific chelators is a potential new strategy for the development of novel anticancer agents. One such chelator, 2-benzoylpyridine 4-ethyl-3-thiosemicarbazone (Bp4eT), possesses potent and selective anticancer activity (J Med Chem 50:3716-3729, 2007). To elucidate the mechanisms of its potent antitumor activity, Bp4eT was labeled with (14)C. Its efficacy was then compared with the (14)C-labeled iron chelator pyridoxal isonicotinoyl hydrazone (PIH), which exhibits low anticancer activity. The ability of these ligands to permeate the cell membrane and their cellular retention was examined under various conditions using SK-N-MC neuroepithelioma cells. The rate of [(14)C]PIH uptake into cells was significantly (p < 0.001) lower than that of [(14)C]Bp4eT at 37??C, indicating that the increased hydrophilicity of [(14)C]PIH reduced membrane permeability. In contrast, the efflux of [(14)C]PIH was significantly (p < 0.05) higher than that of [(14)C]Bp4eT, leading to increased cellular retention of [(14)C]Bp4eT. In addition, the uptake and release of the (14)C-labeled chelators was not reduced by metabolic inhibitors, indicating that these processes were energy-independent. No significant differences were evident in the uptake of [(14)C]Bp4eT at 37 or 4??C, demonstrating a temperature-independent mechanism. Furthermore, adjusting the pH of the culture medium to model the tumor microenvironment did not affect [(14)C]Bp4eT membrane transport. It can be concluded that [(14)C]Bp4eT more effectively permeated the cell membrane and evaded rapid efflux in contrast to [(14)C]PIH. This property, in part, accounts for the more potent anticancer activity of Bp4eT relative to PIH. %Z FOR Codes: 111205 %0 Journal Article %~ PubMed %A Richardson, Des R %A Lane, Darius J R %A Becker, Erika M %A Huang, Michael L-H %A Whitnall, Megan %A Rahmanto, Yohan Suryo %A Sheftel, Alex D %A Ponka, Prem %T Mitochondrial iron trafficking and the integration of iron metabolism between the mitochondrion and cytosol. %B Proceedings of the National Academy of Sciences of the United States of America %D 2010 %C United States %I National Academy of Sciences %V 107 %N 24 %P 10775-82 %@ 0027-8424 %X The mitochondrion is well known for its key role in energy transduction. However, it is less well appreciated that it is also a focal point of iron metabolism. Iron is needed not only for heme and iron sulfur cluster (ISC)-containing proteins involved in electron transport and oxidative phosphorylation, but also for a wide variety of cytoplasmic and nuclear functions, including DNA synthesis. The mitochondrial pathways involved in the generation of both heme and ISCs have been characterized to some extent. However, little is known concerning the regulation of iron uptake by the mitochondrion and how this is coordinated with iron metabolism in the cytosol and other organelles (e.g., lysosomes). In this article, we discuss the burgeoning field of mitochondrial iron metabolism and trafficking that has recently been stimulated by the discovery of proteins involved in mitochondrial iron storage (mitochondrial ferritin) and transport (mitoferrin-1 and -2). In addition, recent work examining mitochondrial diseases (e.g., Friedreich''s ataxia) has established that communication exists between iron metabolism in the mitochondrion and the cytosol. This finding has revealed the ability of the mitochondrion to modulate whole-cell iron-processing to satisfy its own requirements for the crucial processes of heme and ISC synthesis. Knowledge of mitochondrial iron-processing pathways and the interaction between organelles and the cytosol could revolutionize the investigation of iron metabolism. %Z FOR Codes: 110202 60104 %0 Journal Article %~ PubMed %A Jansson, Patric J %A Sharpe, Philip C %A Bernhardt, Paul V %A Richardson, Des R %T Novel Thiosemicarbazones of the ApT and DpT Series and Their Copper Complexes: Identification of Pronounced Redox Activity and Characterization of Their Antitumor Activity. %B Journal of medicinal chemistry %D 2010 %C United States %I American Chemical Society %V 53 %N 15 %P 5759-69 %@ 0022-2623 %X The novel chelators 2-acetylpyridine-4,4-dimethyl-3-thiosemicarbazone (HAp44mT) and di-2-pyridylketone-4,4-dimethyl-3-thiosemicarbazone (HDp44mT) have been examined to elucidate the structure-activity relationships necessary to form copper (Cu) complexes with pronounced antitumor activity. Electrochemical studies demonstrated that the Cu complexes of these ligands had lower redox potentials than their iron complexes. Moreover, the Cu complexes where the ligand/metal ratio was 1:1 rather than 2:1 had significantly higher intracellular oxidative properties and antitumor efficacy. Interestingly, the 2:1 complex was shown to dissociate to give significant amounts of the 1:1 complex that could be the major cytotoxic effector. Both types of Cu complex showed significantly more antiproliferative activity than the ligand alone. We also demonstrate the importance of the inductive effects of substituents on the carbonyl group of the parent ketone, which influence the Cu(II/I) redox potentials because of their proximity to the metal center. The structure-activity relationships described are important for the design of potent thiosemicarbazone Cu complexes. %Z FOR Codes: 111204 %0 Journal Article %~ Isi %A Singleton, W. C. J. %A McInnes, K. T. %A Cater, M. A. %A Winnall, W. R. %A McKirdy, R. %A Yu, Y. %A Taylor, P. E. %A Ke, B. X. %A Richardson, D. R. %A Mercer, J. F. B. %A La Fontaine, S. %T Role of Glutaredoxin1 and Glutathione in Regulating the Activity of the Copper-transporting P-type ATPases, ATP7A and ATP7B %B Journal of Biological Chemistry %D 2010 %C United States %I American Society for Biochemistry and Molecular Biology, Inc. %V 285 %N %P 27111-27121 %@ 0021-9258 %X %Z FOR Codes: 30401 %0 Journal Article %~ PubMed %A Saletta, Federica %A Rahmanto, Yohan Suryo %A Richardson, Des R %T The Translational Regulator eIF3a: The Tricky eIF3 Subunit! %B Biochimica et biophysica acta %D 2010 %C Netherlands %I Elsevier BV %V 1806 %N 2 %P 275-86 %@ 0006-3002 %X Regulation of gene expression is a fundamental step in cellular physiology as abnormalities in this process may lead to de-regulated growth and cancer. Translation of mRNA is mainly regulated at the rate-limiting initiation step, where many eukaryotic initiation factors (eIFs) are involved. The largest and most complex initiation factor is eIF3 which plays a role in translational regulation, cell growth and cancer. The largest subunit of eIF3 is eIF3a, although it is not required for the general function of eIF3 in translation initiation. However, eIF3a may play a role as a regulator of a subset of mRNAs and has been demonstrated to regulate the expression of p27(kip1), tyrosinated ??-tubulin and ribonucleotide reductase M2 subunit. These molecules have a pivotal role in the regulation of the cell cycle. Moreover, the eIF3a mRNA is ubiquitously expressed in all tissues at different levels and is found elevated in a number of cancer types. eIF3a can modulate the cell cycle and may be a translational regulator for proteins important for entrance into S phase. The expression of eIF3a is decreased in differentiated cells in culture and the suppression of eIF3a expression can reverse the malignant phenotype and change the sensitivity of cells to cell cycle modulators. However, the role of eIF3a in cancer is still unclear. In fact, some studies have identified eIF3a to be involved in cancer development, while other results indicate that it could provide protection against evolution into higher malignancy. Together, these findings highlight the "tricky" and interesting nature of eIF3a. %Z FOR Codes: 60111 60103 111201 %0 Journal Article %~ PubMed %A Richardson, Des %A Huang, Michael %A Whitnall, Megan %A Becker, Erika %A Ponka, Prem %A Suryo Rahmanto, Yohan %T The ins and outs of mitochondrial iron-loading: the metabolic defect in Friedreich's ataxia. %B Journal of molecular medicine (Berlin, Germany) %D 2010 %C Germany %I Springer %V 88 %N 4 %P 323-9 %@ 1432-1440 %X Friedreich''s ataxia is a cardio- and neurodegenerative disease due to decreased expression of the mitochondrial protein, frataxin. This defect results in mitochondrial iron-overload, and in this review, we discuss the mechanisms that lead to this iron accumulation. Using a conditional knockout mouse model where frataxin is deleted in the heart, it has been shown that this mutation leads to transferrin receptor-1 upregulation, resulting in increased iron uptake from transferrin. There is also marked downregulation of ferritin that is required for iron storage and decreased expression of the iron exporter, ferroportin 1, leading to decreased cellular iron efflux. The increased mitochondrial iron uptake is facilitated by upregulation of the mitochondrial iron transporter, mitoferrin 2. This stimulation of iron uptake probably attempts to rescue the deficit in mitochondrial iron metabolism that is due to downregulation of mitochondrial iron utilization, namely, heme and iron-sulfur cluster (ISC) synthesis and also iron storage (mitochondrial ferritin). The resultant decrease in heme and ISC synthesis means heme and ISCs are not exiting the mitochondrion for cytosolic use. Hence, increased mitochondrial iron uptake coupled with decreased utilization and release leads to mitochondrial iron-loading. More generally, disturbance of mitochondrial iron utilization in other diseases probably also results in similar compensatory alterations. %Z FOR Codes: 110201 110202 110199 %0 Journal Article %~ PubMed %A Jansson, Patric J %A Hawkins, Clare L %A Lovejoy, David B %A Richardson, Des R %T The iron complex of Dp44mT is redox-active and induces hydroxyl radical formation: An EPR study. %B Journal of inorganic biochemistry %D 2010 %C United States %I Elsevier Inc. %V 104 %N 11 %P 1224-8 %@ 0162-0134 %X Iron chelation therapy was initially designed to alleviate the toxic effects of excess iron evident in iron-overload diseases. However, some iron chelator-metal complexes have also gained interest due to their high redox activity and toxicological properties that have potential for cancer chemotherapy. This communication addresses the conflicting results published recently on the ability of the iron chelator, Dp44mT, to induce hydroxyl radical formation upon complexation with iron (B.B. Hasinoff and D. Patel, J Inorg. Biochem.103 (2009), 1093-1101). This previous study used EPR spin-trapping to show that Dp44mT-iron complexes were not able to generate hydroxyl radicals. Here, we demonstrate the opposite by using the same technique under very similar conditions to show the Dp44mT-iron complex is indeed redox-active and induces hydroxyl radical formation. This was studied directly in an iron(II)/H(2)O(2) reaction system or using a reducing iron(III)/ascorbate system implementing several different buffers at pH 7.4. The demonstration by EPR that the Dp44mT-iron complex is redox-active confirms our previous studies using cyclic voltammetry, ascorbate oxidation, benzoate hydroxylation and a plasmid DNA strand-break assay. We discuss the relevance of the redox activity to the biological effects of Dp44mT. %Z FOR Codes: 30201 60101 %0 Journal Article %~ PubMed %A Richardson, Des R %A Kalinowski, Danuta S %A Richardson, Vera %A Sharpe, Philip C %A Lovejoy, David B %A Islam, Mohammad %A Bernhardt, Paul V %T 2-Acetylpyridine Thiosemicarbazones are Potent Iron Chelators and Antiproliferative Agents: Redox Activity, Iron Complexation and Characterization of their Antitumor Activity. %B Journal of medicinal chemistry %D 2009 %C United States %I American Chemical Society %V 52 %N 5 %P 1459-70 %@ 0022-2623 %X Through systematic structure-activity studies of the 2-benzoylpyridine thiosemicarbazone (HBpT), 2-(3-nitrobenzoyl)pyridine thiosemicarbazone (HNBpT) and dipyridylketone thiosemicarbazone (HDpT) series of iron (Fe) chelators, we identified structural features necessary to form Fe complexes with potent anticancer activity (J. Med. Chem. 2007, 50, 3716-3729). In this investigation, we generated the related 2-acetylpyridine thiosemicarbazone (HApT) analogues to examine the influence of the methyl group at the imine carbon. Four of the six HApT chelators had potent antitumor activity (IC(50): 0.001-0.002 microM) and Fe chelation efficacy that was similar to the most effective HBpT and HDpT ligands. The HApT Fe complexes had the lowest Fe(III/II) redox potentials of any thiosemicarbazone series we have generated. This property, in combination with their ability to effectively chelate cellular Fe, make the HApT series one of the most potent antiproliferative agents developed by our group. %Z FOR Codes: 110199 %0 Journal Article %~ PubMed %A Bohlken, A %A Cheung, B B %A Bell, J L %A Koach, J %A Smith, S %A Sekyere, E %A Thomas, W %A Norris, M %A Haber, M %A Lovejoy, D B %A Richardson, D R %A Marshall, G M %T ATP7A is a novel target of retinoic acid receptor beta2 in neuroblastoma cells. %B British Journal of Cancer %D 2009 %C United Kingdom %I Nature Publishing Group %V 100 %N 1 %P 96-105 %@ 0007-0920 %X Increased retinoic acid receptor beta (RARbeta(2)) gene expression is a hallmark of cancer cell responsiveness to retinoid anticancer effects. Moreover, low basal or induced RARbeta(2) expression is a common feature of many human cancers, suggesting that RARbeta(2) may act as a tumour suppressor gene in the absence of supplemented retinoid. We have previously shown that low RARbeta(2) expression is a feature of advanced neuroblastoma. Here, we demonstrate that the ABC domain of the RARbeta(2) protein alone was sufficient for the growth inhibitory effects of RARbeta(2) on neuroblastoma cells. ATP7A, the copper efflux pump, is a retinoid-responsive gene, was upregulated by ectopic overexpression of RARbeta(2). The ectopic overexpression of the RARbeta(2) ABC domain was sufficient to induce ATP7A expression, whereas, RARbeta(2) siRNA blocked the induction of ATP7A expression in retinoid-treated neuroblastoma cells. Forced downregulation of ATP7A reduced copper efflux and increased viability of retinoid-treated neuroblastoma cells. Copper supplementation enhanced cell growth and reduced retinoid-responsiveness, whereas copper chelation reduced the viability and proliferative capacity. Taken together, our data demonstrates ATP7A expression is regulated by retinoic acid receptor beta and it has effects on intracellular copper levels, revealing a link between the anticancer action of retinoids and copper metabolism. %Z FOR Codes: 111201 %0 Journal Article %~ Isi %A Noulsri, E. %A Richardson, D. R. %A Lerdwana, S. %A Fucharoen, S. %A Yamagishi, T. %A Kalinowski, D. S. %A Pattanapanyasat, K. %T Antitumor activity and mechanism of action of the iron chelator, Dp44mT, against leukemic cells %B American Journal of Hematology %D 2009 %C United States %I John Wiley & Sons, Inc. %V 84 %N 3 %P 170-176 %@ 0361-8609 %X %Z FOR Codes: 30405 %0 Journal Article %~ PubMed %A Huang, Michael Li-Hsuan %A Becker, Erika M %A Whitnall, Megan %A Rahmanto, Yohan Suryo %A Ponka, Prem %A Richardson, Des R %T Elucidation of the mechanism of mitochondrial iron loading in Friedreich's ataxia by analysis of a mouse mutant. %B Proceedings of the National Academy of Sciences of the United States of America %D 2009 %C United States %I National Academy of Sciences %V 106 %N 38 %P 16381-16386 %@ 0027-8424 %X We used the muscle creatine kinase (MCK) conditional frataxin knockout mouse to elucidate how frataxin deficiency alters iron metabolism. This is of significance because frataxin deficiency leads to Friedreich''s ataxia, a disease marked by neurologic and cardiologic degeneration. Using cardiac tissues, we demonstrate that frataxin deficiency leads to down-regulation of key molecules involved in 3 mitochondrial utilization pathways: iron-sulfur cluster (ISC) synthesis (iron-sulfur cluster scaffold protein1/2 and the cysteine desulferase Nfs1), mitochondrial iron storage (mitochondrial ferritin), and heme synthesis (5-aminolevulinate dehydratase, coproporphyrinogen oxidase, hydroxymethylbilane synthase, uroporphyrinogen III synthase, and ferrochelatase). This marked decrease in mitochondrial iron utilization and resultant reduced release of heme and ISC from the mitochondrion could contribute to the excessive mitochondrial iron observed. This effect is compounded by increased iron availability for mitochondrial uptake through (i) transferrin receptor1 up-regulation, increasing iron uptake from transferrin; (ii) decreased ferroportin1 expression, limiting iron export; (iii) increased expression of the heme catabolism enzyme heme oxygenase1 and down-regulation of ferritin-H and -L, both likely leading to increased "free iron" for mitochondrial uptake; and (iv) increased expression of the mammalian exocyst protein Sec15l1 and the mitochondrial iron importer mitoferrin-2 (Mfrn2), which facilitate cellular iron uptake and mitochondrial iron influx, respectively. Our results enable the construction of a model explaining the cytosolic iron deficiency and mitochondrial iron loading in the absence of frataxin, which is important for understanding the pathogenesis of Friedreich''s ataxia. %Z FOR Codes: 110201 %0 Journal Article %~ PubMed %A Suryo, Rahmanto Y %A Richardson, D R %T Generation and characterization of transgenic mice hyper-expressing melanoma tumour antigen p97 (Melanotransferrin): No overt alteration in phenotype. %B Biochimica et biophysica acta %D 2009 %C Melbourne, Vic %I Council on the Ageing %V 1793 %N 0 %P 1210-7 %@ 0006-3002 %X Melanotransferrin (MTf) is a transferrin homologue that binds iron (Fe) through a high affinity Fe-binding site. MTf has been implicated in diverse processes, e.g., iron metabolism, plasminogen activation, eosinophil differentiation and cancer cell migration, proliferation and tumourigenesis. Our previous studies using a knockout mouse demonstrated that MTf does not have an essential function in Fe metabolism (E.O. Sekyere, L.L. Dunn, Y.S. Rahmanto, D.R. Richardson, Role of melanotransferrin in iron metabolism: studies using targeted gene disruption in vivo, Blood 107 (2006) 2599-2601). However, it does play a role in melanoma cell proliferation and tumourigenesis. In this investigation, we report generation and characterization of transgenic mice bearing the MTf gene (MTf(Tg)) produced via lentiviral delivery. In MTf(Tg) mice, MTf mRNA and protein were hyper-expressed in tissues compared to control mice. These animals exhibited no gross morphological, histological, nor Fe status changes. The MTf(Tg) mice were also born in accordance with classical Mendelian ratios. However, hyper-expression of MTf leads to a mild, but significant decrease in erythrocyte count. This animal provides a novel MTf hyper-expression transgenic model for further investigating the biological function(s) of MTf. %Z FOR Codes: 111201 111201 %0 Journal Article %~ PubMed %A Rosemary Siafakas, A %A Richardson, Des R %T Growth arrest and DNA damage-45 alpha (GADD45alpha). %B The international journal of biochemistry & cell biology %D 2009 %C United Kingdom %I Pergamon %V 41 %N 0 %P 986-9 %@ 1357-2725 %X Regulation of cell cycle and growth is integral for cell survival. The intricate mechanisms that control proliferation and cell cycle are numerous. The growth arrest and DNA damage (GADD)-inducible gene family is often up-regulated in response to various environmental stresses and drug therapies. GADD45alpha was the first stress-inducible gene determined to be up-regulated by p53 and is also a target for the p53 homologues, p63 and p73. When GADD45alpha is deleted or repressed, cells show uncontrolled proliferation. Furthermore, decreased GADD45alpha expression is also considered a survival mechanism, as cancer cells without this control can evade the apoptotic pathway leading to increased tumourigenesis. Drug therapies can act to directly or indirectly up-regulate GADD45alpha and promote apoptosis. As GADD45alpha is an essential component of many metabolic pathways that control proliferating cancer cells, it presents itself as an emerging drug target worthy of further investigation. %Z FOR Codes: 1101 %0 Journal Article %~ PubMed %A Stariat, Jn %A Kovarkov, Petra %A Klimes, Jir %A Lovejoy, David B %A Kalinowski, Danuta S %A Richardson, Des R %T HPLC methods for determination of two novel thiosemicarbazone anti-cancer drugs (N4mT and Dp44mT) in plasma and their application to in vitro plasma stability of these agents. %B Journal of chromatography. B, Analytical technologies in the biomedical and life sciences %D 2009 %C Netherlands %I Elsevier BV %V 877 %N 3 %P 316-322 %@ 1570-0232 %X The aim of this study was to develop and validate HPLC methods for the determination in plasma of two novel thiosemicarbazone anti-tumour drugs developed in our laboratories (Dp44mT and N4mT). The appropriate separations were achieved using a HS F5 HPLC column with the mobile phase composed of a mixture of either acetate buffer/EDTA or EDTA and acetonitrile (62:38 and 50:50, v/v, respectively). The plasma samples were pretreated with SPE (phenyl and C18, respectively). Furthermore, these methods were successfully applied to in vitro plasma stability experiments. The investigation has clearly shown that both thiosemicarbazones are markedly more stable in plasma than their aroylhydrazone forerunners. %Z FOR Codes: 110899 %0 Journal Article %~ PubMed %A Peslova, Gabriela %A Petrak, Jiri %A Kuzelova, Katerina %A Hrdy, Ivan %A Halada, Petr %A Kuchel, Philip W %A Soe-Lin, Shan %A Ponka, Prem %A Sutak, Robert %A Becker, Erika %A Huang, Michael Li-Hsuan %A Suryo Rahmanto, Yohan %A Richardson, Des R %A Vyoral, Daniel %T Hepcidin, the hormone of iron metabolism, is bound specifically to {alpha}-2-macroglobulin in blood. %B Blood %D 2009 %C United States %I American Society of Hematology %V 113 %N 0 %P 6225-36 %@ 1528-0020 %X Hepcidin is a major regulator of iron metabolism. Hepcidin-based therapeutics/diagnostics could play roles in hematology in the future, and thus, hepcidin transport is crucial to understand. In this study, we identify alpha2-macroglobulin (alpha2-M) as the specific hepcidin-binding molecule in blood. Interaction of 125I-hepcidin with alpha2-M was identified using fractionation of plasma proteins followed by native gradient polyacrylamide gel electrophoresis and mass spectrometry. Hepcidin binding to nonactivated alpha2-M displays high affinity (Kd 177 +/- 27 nM), whereas hepcidin binding to albumin was nonspecific and displayed nonsaturable kinetics. Surprisingly, the interaction of hepcidin with activated alpha2-M exhibited a classical sigmoidal binding curve demonstrating cooperative binding of 4 high-affinity (Kd 0.3 microM) hepcidin-binding sites. This property probably enables efficient sequestration of hepcidin and its subsequent release or inactivation that may be important for its effector functions. Because alpha2-M rapidly targets ligands to cells via receptor-mediated endocytosis, the binding of hepcidin to alpha2-M may influence its functions. In fact, the alpha2-M-hepcidin complex decreased ferroportin expression in J774 cells more effectively than hepcidin alone. The demonstration that alpha2-M is the hepcidin transporter could lead to better understanding of hepcidin physiology, methods for its sensitive measurement and the development of novel drugs for the treatment of iron-related diseases. %Z FOR Codes: 30401 %0 Journal Article %~ PubMed %A Saletta, Federica %A Suryo Rahmanto, Yohan %A Noulsri, Egarit %A Richardson, Des R %T Iron Chelator-Mediated Alterations in Gene Expression: Identification of Novel Iron-Regulated Molecules that are Molecular Targets of HIF-1{alpha} and p53. %B Molecular pharmacology %D 2009 %C United States %I American Society for Pharmacology and Experimental Therapeutics %V 77 %N 0 %P 443-58 %@ 1521-0111 %X Iron deficiency affects 500 million people, yet the molecular role of iron in gene expression remains poorly characterized. In addition, the alterations in global gene expression after iron chelation remain unclear and are important to assess for understanding the molecular pathology of iron deficiency and the biological effects of chelators. Considering this, we assessed the effect on whole genome gene expression of two iron chelators (desferrioxamine and 2-hydroxy-1-napthylaldehyde isonicotinoyl hydrazone) that have markedly different permeability properties. Sixteen genes were significantly regulated by both ligands, whereas a further 50 genes were significantly regulated by either compound. Apart from iron-mediated regulation of expression via hypoxia inducible factor-1 alpha, it was noteworthy that the transcription factor p53 was also involved in iron-regulated gene expression. Examining 16 genes regulated by both chelators in normal and neoplastic cells, five genes (APP, GDF15, CITED2, EGR1, and PNRC1) were significantly differentially expressed between the cell types. In view of their functions in tumor suppression, proliferation, and apoptosis, these findings are important for understanding the selective antiproliferative effects of chelators against neoplastic cells. Most of the genes identified have not been described previously to be iron-regulated and are important for understanding the molecular and cellular effects of iron depletion. %Z FOR Codes: 60199 110202 110105 %0 Journal Article %~ PubMed %A Bernhardt, Paul V %A Sharpe, Philip C %A Islam, Mohammad %A Lovejoy, David B %A Kalinowski, Danuta S %A Richardson, Des R %T Iron Chelators of the Dipyridylketone Thiosemicarbazone Class: Precomplexation and Transmetalation Effects on Anticancer Activity. %B Journal of medicinal chemistry %D 2009 %C United States %I American Chemical Society %V 52 %N 2 %P 407-15 %@ 0022-2623 %X We previously reported a series of di-2-pyridylketone thiosemicarbazone (HDpT) chelators that showed marked and selective antitumor activity (Whitnall, M.; et al. Proc. Natl. Acad. Sci. U.S.A. 2006, 103, 14901-14906). To further understand their biological efficacy, we report the characterization and activity of their Mn(II), Co(III), Ni(II), Cu(II), and Zn(II) complexes. The X-ray crystal structures of four divalent (Mn, Ni, Cu, and Zn) and one trivalent (Fe) complexes are reported. Electrochemistry shows the Fe(III/II) and Cu(II/I) potentials of the complexes may be redox-active within cells. Stability constants were also determined for the Mn(II), Ni(II), Cu(II), and Zn(II) complexes. All divalent complexes underwent transmetalation upon encountering Fe(II), to form low spin ferrous complexes. Importantly, the divalent Mn(II), Ni(II), Cu(II), and Zn(II) complexes of the HDpT analogues are equally active in preventing proliferation as their ligands, suggesting the complexes act as lipophilic vehicles facilitating intracellular delivery of the free ligand upon metal dissociation. %Z FOR Codes: 111204 110199 %0 Journal Article %~ Isi %A Sheftel, A. D. %A Richardson, D. R. %A Prchal, J. %A Ponka, P. %T Mitochondrial Iron Metabolism and Sideroblastic Anemia %B Acta Haematologica %D 2009 %C Switzerland %I S. Karger AG %V 122 %N 2-3 %P 120-133 %@ 0001-5792 %X %Z FOR Codes: 60104 %0 Journal Article %~ PubMed %A Assinder, Stephen J %A Dong, Qihan %A Mangs, Helena %A Richardson, Des R %T Pharmacological Targeting of the Integrated AKT, PTEN and TGF-{beta} Pathways in Prostate Cancer. %B Molecular pharmacology %D 2009 %C United States %I Amer Soc Pharmacology Experimental Therapeutics %V 75 %N 3 %P 429-36 %@ 0026-895X %X Prostate cancer is a highly heterogenous disease in which a patient-tailored care program is much desired. Central to this goal is the development of novel targeted pharmacological interventions. To develop these treatment strategies, an understanding of the integration of cellular pathways involved in both tumorigenesis and tumor suppression is crucial. Of further interest are the events elicited by drug treatments that exploit the underlying molecular pathology in cancer. This review briefly describes the evidence that suggests integration of three established pathways: the tumorigenic phosphoinositide 3-kinase/protein kinase B (AKT) pathway, the tumor suppressive phosphatase and tensin homolog deleted on chromosome 10 pathway, and the tumor suppressive transforming growth factor-beta pathway. More importantly, we discuss novel pharmaceutical agents that target key points of integration in these three pathways. These new therapeutic strategies include the use of agents that target iron to inhibit proliferation via multiple mechanisms and suppression of AKT by cytosolic phospholipase A(2)-alpha inhibitors. %Z FOR Codes: 111201 %0 Journal Article %~ PubMed %A Assinder, Stephen J %A Dong, Qihan %A Kovacevic, Zaklina %A Richardson, Des R %T The TGF-beta, PI3K/Akt and PTEN pathways: established and proposed biochemical integration in prostate cancer. %B Biochemical Journal %D 2009 %C United Kingdom %I Portland Press Ltd. %V 417 %N 2 %P 411-421 %@ 1470-8728 %X A key to the development of improved pharmacological treatment strategies for cancer is an understanding of the integration of biochemical pathways involved in both tumorigenesis and cancer suppression. Furthermore, genetic markers that may predict the outcome of targeted pharmacological intervention in an individual are central to patient-focused treatment regimens rather than the traditional ''one size fits all'' approach. Prostate cancer is a highly heterogeneous disease in which a patient-tailored care program is a holy grail. This review will describe the evidence that demonstrates the integration of three established pathways: the tumour-suppressive TGF-beta (transforming growth factor-beta) pathway, the tumorigenic PI3K/Akt (phosphoinositide 3-kinase/protein kinase B) pathway and the tumour-suppressive PTEN (phosphatase and tensin homologue deleted on chromosome 10) pathway. It will discuss gene polymorphisms and somatic mutations in relevant genes and highlight novel pharmaceutical agents that target key points in these integrated pathways. %Z FOR Codes: 111201 %0 Journal Article %~ PubMed %A Mlad?nka, Premysl %A Kalinowski, Danuta S %A Haskova, Pavlna %A Bobrovov, Zuzana %A Hrdina, Radomr %A Sim?nek, Toms %A Nachtigal, Petr %A Semeck, Vladim?r %A Vvrov, Jaroslava %A Holeckova, Magdalna %A Palicka, Vladimir %A Mazurov, Yvona %A Jansson, Patric J %A Richardson, Des R %T The novel iron chelator, 2-pyridylcarboxaldehyde 2-thiophenecarboxyl hydrazone, reduces catecholamine-mediated myocardial toxicity. %B Chemical research in toxicology %D 2009 %C United States %I American Chemical Society %V 22 %N 1 %P 208-217 %@ 1520-5010 %X Iron (Fe) chelators are used clinically for the treatment of Fe overload disease. Iron also plays a role in the pathology of many other conditions, and these potentially include the cardiotoxicity induced by catecholamines such as isoprenaline (ISO). The current study examined the potential of Fe chelators to prevent ISO cardiotoxicity. This was done as like other catecholamines, ISO contains the classical catechol moiety that binds Fe and may form redox-active and cytotoxic Fe complexes. Studies in vitro used the cardiomyocyte cell line, H9c2, which was treated with ISO in the presence or absence of the chelator, desferrioxamine (DFO), or the lipophilic ligand, 2-pyridylcarboxaldehyde 2-thiophenecarboxyl hydrazone (PCTH). Both of these chelators were not cardiotoxic and significantly reduced ISO cardiotoxicity in vitro. However, PCTH was far more effective than DFO, with the latter showing activity only at a high, clinically unachievable concentration. Further studies in vitro showed that interaction of ISO with Fe(II)/(III) did not increase cytotoxic radical generation, suggesting that this mechanism was not involved. Studies in vivo were initiated using rats pretreated intravenously with DFO or PCTH before subcutaneous administration of ISO (100 mg/kg). DFO at a clinically used dose (50 mg/kg) failed to reduce catecholamine cardiotoxicity, while PCTH at an equimolar dose totally prevented catecholamine-induced mortality and reduced cardiotoxicity. This study demonstrates that PCTH reduced ISO-induced cardiotoxicity in vitro and in vivo, demonstrating that Fe plays a role, in part, in the pathology observed. %Z FOR Codes: 110899 %0 Journal Article %~ PubMed %A Yu, Yu %A Kalinowski, Danuta S %A Kovacevic, Zaklina %A Siafakas, Aritee R %A Jansson, Patric J %A Stefani, Christian %A Lovejoy, David B %A Sharpe, Philip C %A Bernhardt, Paul V %A Richardson, Des R %T Thiosemicarbazones from the Old to New: Iron Chelators That Are More Than Just Ribonucleotide Reductase Inhibitors. %B Journal of medicinal chemistry %D 2009 %C United States %I American Chemical Society %V 52 %N 17 %P 5271-94 %@ 0022-2623 %X %Z FOR Codes: 30499 %0 Journal Article %~ PubMed %A Kalinowski, Danuta S %A Quach, Patricia %A Richardson, Des R %T Thiosemicarbazones: the new wave in cancer treatment. %B Future Medicinal Chemistry %D 2009 %C United Kingdom %I Future Science Ltd. %V 1 %N 6 %P 1143-1151 %@ 1756-8927 %X The anticancer effects of thiosemicarbazones were once solely attributed to the inhibition of ribonucleotide reductase, an enzyme involved in the rate-limiting step of DNA synthesis. However, the mechanism behind this inhibition was initially not described. The ability of thiosemicarbazones to chelate metal ions has now been recognized as a major factor in their antiproliferative effects. This mini-review discusses current advances of an emerging ''new wave'' of thiosemicarbazones as potent anticancer agents, describing recent insights into their mechanism of action. The redox activity of Fe-thiosemicarbazone complexes is critical in their anticancer activity, resulting in oxidative damage and the inhibition of ribonucleotide reductase. In vivo analysis indicates that some thiosemicarbazones show potential as chemotherapeutic agents. %Z FOR Codes: 30401 %0 Journal Article %~ PubMed %A Long, Shaojun %A Jirk?, Milan %A Mach, Jan %A Ginger, Michael L %A Sutak, Robert %A Richardson, Des %A Tachezy, Jan %A Luke?, Julius %T Ancestral roles of eukaryotic frataxin: mitochondrial frataxin function and heterologous expression of hydrogenosomal Trichomonas homologues in trypanosomes. %B Molecular microbiology %D 2008 %C United Kingdom %I Wiley-Blackwell %V 69 %N 1 %P 94-109 %@ 0950-382X %X Frataxin is a small conserved mitochondrial protein; in humans, mutations affecting frataxin expression or function result in Friedreich''s ataxia. Much of the current understanding of frataxin function comes from informative studies with yeast models, but considerable debates remain with regard to the primary functions of this ubiquitous protein. We exploit the tractable reverse genetics of Trypanosoma brucei in order to specifically consider the importance of frataxin in an early branching lineage. Using inducible RNAi, we show that frataxin is essential in T. brucei and that its loss results in reduced activity of the marker Fe-S cluster-containing enzyme aconitase in both the mitochondrion and cytosol. Activities of mitochondrial succinate dehydrogenase and fumarase also decreased, but the concentration of reactive oxygen species increased. Trypanosomes lacking frataxin also exhibited a low mitochondrial membrane potential and reduced oxygen consumption. Crucially, however, iron did not accumulate in frataxin-depleted mitochondria, and as T. brucei frataxin does not form large complexes, it suggests that it plays no role in iron storage. Interestingly, RNAi phenotypes were ameliorated by expression of frataxin homologues from hydrogenosomes of another divergent protist Trichomonas vaginalis. Collectively, the data suggest trypanosome frataxin functions primarily only in Fe-S cluster biogenesis and protection from reactive oxygen species. %Z FOR Codes: 110899 %0 Journal Article %~ PubMed %A Farnaud, Sebastien %A Amini, Maryam %A Rapisarda, Chiara %A Cammack, Richard %A Bui, Tam %A Drake, Alex %A Evans, Robert W %A Rahmanto, Yohan Suryo %A Richardson, Des R %T Biochemical and spectroscopic studies of human melanotransferrin (MTf): electron-paramagnetic resonance evidence for a difference between the iron-binding site of MTf and other transferrins. %B The international journal of biochemistry & cell biology %D 2008 %C United Kingdom %I Pergamon %V 40 %N 12 %P 2739-45 %@ 1357-2725 %X Melanotransferrin (MTf) is a member of the transferrin (Tf) family of iron (Fe)-binding proteins that was first identified as a cell-surface marker of melanoma. Although MTf has a high-affinity Fe-binding site that is practically identical to that of serum Tf, the protein does not play an essential role in Fe homeostasis and its precise molecular function remains unclear. A Zn(II)-binding motif, distinct from the Fe-binding site, has been proposed in human MTf based on computer modelling studies. However, little is known concerning the interaction of its proposed binding site(s) with metals and the consequences in terms of MTf conformation. For the first time, biochemical and spectroscopic techniques have been used in this study to characterise metal ion-binding to recombinant MTf. Initially, the binding of Fe to MTf was examined using 6M urea gel electrophoresis. Although four different iron-loaded forms were observed with serum Tf, only two forms were found with MTf, the apo-form and the N-monoferric holo-protein, suggesting a single high-affinity site. The presence of a single Fe(III)-binding site was also supported by EPR results which indicated that the Fe(III)-binding characteristics of MTf were unique, but somewhat comparable to the N-lobes of human serum Tf and chicken ovo-Tf. Circular dichroism (CD) analysis indicated that, as for Tf, no changes in secondary structure could be observed upon Fe(III)-binding. The ability of MTf to bind Zn(II) was also investigated using CD which demonstrated that the single high-affinity Fe-binding site was distinct from a potential Zn(II)-binding site. %Z FOR Codes: 110199 110199 %0 Journal Article %~ PubMed %A Richardson, D R %A Kalinowski, D S %A Lau, S %A Jansson, P J %A Lovejoy, D B %T Cancer cell iron metabolism and the development of potent iron chelators as anti-tumour agents. %B Biochimica et biophysica acta %D 2008 %C Netherlands %I Elsevier BV %V 1790 %N 0 %P 702-17 %@ 0006-3002 %X Cancer contributes to 50% of deaths worldwide and new anti-tumour therapeutics with novel mechanisms of actions are essential to develop. Metabolic inhibitors represent an important class of anti-tumour agents and for many years, agents targeting the nutrient folate were developed for the treatment of cancer. This is because of the critical need of this factor for DNA synthesis. Similarly to folate, Fe is an essential cellular nutrient that is critical for DNA synthesis. However, in contrast to folate, there has been limited effort applied to specifically design and develop Fe chelators for the treatment of cancer. Recently, investigations have led to the generation of novel di-2-pyridylketone thiosemicarbazone (DpT) and 2-benzoylpyridine thiosemicarbazone (BpT) group of ligands that demonstrate marked and selective anti-tumour activity in vitro and also in vivo against a wide spectrum of tumours. Indeed, administration of these compounds to mice did not induce whole body Fe-depletion or disturbances in haematological or biochemical indices due to the very low doses required. The mechanism of action of these ligands includes alterations in expression of molecules involved in cell cycle control and metastasis suppression, as well as the generation of redox-active Fe complexes. This review examines the alterations in Fe metabolism in tumour cells and the systematic development of novel aroylhydrazone and thiosemicarbazone Fe chelators for cancer treatment. %Z FOR Codes: 110316 %0 Book Section %A Lovejoy, David %A Yu, Yu %A Richardson, Des %T Chelators as anti-cancer drugs %B Encyclopedia of Cancer %D 2008 %C Germany %I Springer %V %N %P 0 %@ 9783540368472 %E Schwab, Manfred %X %Z FOR Codes: 111299 %0 Journal Article %~ PubMed %A Sutak, Robert %A Lesuisse, Emmanuel %A Tachezy, Jan %A Richardson, Des R %T Crusade for iron: iron uptake in unicellular eukaryotes and its significance for virulence. %B Trends in microbiology %D 2008 %C United Kingdom %I Elsevier %V 16 %N 6 %P 261-8 %@ 0966-842X %X The effective acquisition of iron is a pre-requisite for survival of all organisms, especially parasites that have a high iron requirement. In mammals, iron homeostasis is meticulously regulated; extracellular free iron is essentially unavailable and host iron availability has a crucial role in the host-pathogen relationship. Therefore, pathogens use specialized and effective mechanisms to acquire iron. In this review, we summarize the iron-uptake systems in eukaryotic unicellular organisms with particular focus on the pathogenic species: Candida albicans, Tritrichomonas foetus, Trypanosoma brucei and Leishmania spp. We describe the diversity of their iron-uptake mechanisms and highlight the importance of the process for virulence. %Z FOR Codes: 110199 %0 Journal Article %~ PubMed %A Opletalova??, Veronika %A Kalinowski, Danuta %A Vejsova??, Marcela %A Kunes, Jir %A Pour, Milan %A Jampi??lek, Josef %A Buchta, Vladimi??r %A Richardson, Des %T Identification and Characterization of Thiosemicarbazones with Antifungal and Antitumor Effects: Cellular Iron Chelation Mediating Cytotoxic Activity. %B Chemical research in toxicology %D 2008 %C United States %I American Chemical Society %V 21 %N 9 %P 1878-89 %@ 1520-5010 %X Thiosemicarbazones derived from acetylpyrazines were prepared by condensing an acetylpyrazine or a ring-substituted acetylpyrazine with thiosemicarbazide. Using the same procedure, N, N-dimethylthiosemicarbazones were synthesized from acetylpyrazines and N, N-dimethylthiosemicarbazide. A total of 20 compounds (16 novel) were chemically characterized and then tested for antifungal effects on eight strains of fungi and also for antitumor activity against SK-N-MC neuroepithelioma cells. The most effective compound identified in terms of both antifungal and antitumor activity was N, N-dimethyl-2-(1-pyrazin-2-ylethylidene)hydrazinecarbothioamide (5a). The mechanism of action of this and its related thiosemicarbazones was due, at least in part, to its ability to act as a tridentate ligand that binds metal ions. This was deduced from preparation of the related thiosemicarbazones [acetophenone thiosemicarbazone (6) and acetophenone N, N-dimethylthiosemicarbazone (7)] that do not possess a coordinating ring-N, which plays a vital role in metal ion chelation. Furthermore, 5a and several other thiosemicarbazones that showed high antiproliferative activity were demonstrated to have marked iron (Fe) chelation efficacy. In fact, these agents were highly effective at mobilizing (59)Fe from prelabeled SK-N-MC cells and preventing (59)Fe uptake from the serum Fe transport protein, transferrin. In contrast, compounds 6 and 7 that do not possess a tridentate metal-binding site showed little activity. Further studies examining ascorbate oxidation demonstrated that the Fe complexes of the most effective compounds were redox-inactive. Thus, in contrast to other thiosemicarbazones with potent antiproliferative activity, Fe chelation and mobilization rather than free radical generation played a significant role in the cytotoxic effects of the current ligands. %Z FOR Codes: 110107 %0 Journal Article %~ PubMed %A Lim, Chai K %A Kalinowski, Danuta S %A Richardson, Des R %T Protection Against Hydrogen Peroxide-Mediated Cytotoxicity in Friedreich's Ataxia Fibroblasts using Novel Iron Chelators of the PCIH Class. %B Molecular pharmacology %D 2008 %C United States %I Amer Soc Pharmacology Experimental Therapeutics %V 74 %N 1 %P 225-35 %@ 0026-895X %X Iron-loading diseases remain an important problem because of the toxicity of iron-catalyzed redox reactions. Iron loading occurs in the mitochondria of Friedreich''s ataxia (FA) patients and may play a role in its pathogenesis. This suggests that iron chelation therapy could be useful. We developed previously the lipophilic iron chelators known as the 2-pyridylcarboxaldehyde isonicotinoyl hydrazone (PCIH) ligands and identified 2-pyridylcarboxaldehyde 2-thiophenecarboxyl hydrazone (PCTH) as the most promising analog. Hence, this study assessed the efficacy of PCTH and other PCIH analogs compared with various chelators, including deferiprone and desferrioxamine (DFO). Age- and sex-matched control and FA fibroblasts were preincubated with iron chelators and subsequently challenged with 50 microM H2O2 for up to 24 h. The current study demonstrates an interesting structure-activity relationship among the closely related PCIH series of ligands, with only PCTH being highly effective at preventing H2O2-induced cytotoxicity. PCTH increased FA fibroblast cell viability by up to 70%, whereas DFO rescued viability by 1 to 5% only. Hence, PCTH, which was well tolerated by cells was far more effective than DFO at preventing oxidative stress. It is noteworthy that kinetic studies demonstrated PCTH to rapidly penetrate cells to induce 59Fe efflux, whereas DFO, PCIH, 2-pyridylcarboxaldehyde benzoyl hydrazone, and 2-pyridylcarboxaldehyde m-bromobenzoyl hydrazone were far slower, indicating it is the rate of chelator permeation that is crucial for protection against H2O2. In addition, PCTH was found to be as effective as or more effective than conventional radical scavengers or the antioxidant idebenone (which has undergone clinical trials) at protecting cells against H2O2-mediated cytotoxicity. These findings further indicate the potential of PCTH for treatment of iron overload. %Z FOR Codes: 110316 %0 Journal Article %~ PubMed %A Sutak, Robert %A Xu, Xiangcong %A Whitnall, Megan %A Kashem, Mohammed Abul %A Vyoral, Daniel %A Richardson, Des R %T Proteomic analysis of hearts from frataxin knockout mice: marked rearrangement of energy metabolism, a response to cellular stress and altered expression of proteins involved in cell structure, motility and metabolism. %B Proteomics %D 2008 %C Germany %I Wiley - VCH Verlag GmbH & Co. KGaA %V 8 %N 8 %P 1731-1741 %@ 1615-9853 %X A frequent cause of death in Friedreich''s ataxia patients is cardiomyopathy, but the molecular alterations underlying this condition are unknown. We performed 2-DE to characterize the changes in protein expression of hearts using the muscle creatine kinase frataxin conditional knockout (KO) mouse. Pronounced changes in protein expression profile were observed in 9 week-old KO mice with severe cardiomyopathy. In contrast, only several proteins showed altered expression in asymptomatic 4 week-old KO mice. In hearts from frataxin KO mice, components of the iron-dependent complex-I and -II of the mitochondrial electron transport chain and enzymes involved in ATP homeostasis (creatine kinase, adenylate kinase) displayed decreased expression. Interestingly, the KO hearts exhibited increased expression of enzymes involved in the citric acid cycle, catabolism of branched-chain amino acids, ketone body utilization and pyruvate decarboxylation. This constitutes evidence of metabolic compensation due to decreased expression of electron transport proteins. There was also pronounced up-regulation of proteins involved in stress protection, such as a variety of chaperones, as well as altered expression of proteins involved in cellular structure, motility and general metabolism. This is the first report of the molecular changes at the protein level which could be involved in the cardiomyopathy of the frataxin KO mouse. %Z FOR Codes: 110199 %0 Journal Article %~ PubMed %A Lim, Su Yin %A Raftery, Mark %A Cai, Hong %A Hsu, Kenneth %A Yan, Wei Xing %A Hseih, Hsiao-Ling %A Watts, Ralph N %A Richardson, Des %A Thomas, Shane %A Perry, Michael %A Geczy, Carolyn L %T S-nitrosylated S100A8: novel anti-inflammatory properties. %B Journal of Immunology %D 2008 %C United States %I American Association of Immunologists %V 181 %N 8 %P 5627-5636 %@ 0022-1767 %X S100A8 and S100A9, highly expressed by neutrophils, activated macrophages, and microvascular endothelial cells, are secreted during inflammatory processes. Our earlier studies showed S100A8 to be an avid scavenger of oxidants, and, together with its dependence on IL-10 for expression in macrophages, we postulated that this protein has a protective role. S-nitrosylation is an important posttranslational modification that regulates NO transport, cell signaling, and homeostasis. Relatively few proteins are targets of S-nitrosylation. To date, no inflammation-associated proteins with NO-shuttling capacity have been identified. We used HPLC and mass spectrometry to show that S100A8 and S100A9 were readily S-nitrosylated by NO donors. S-nitrosylated S100A8 (S100A8-SNO) was the preferred nitrosylated product. No S-nitrosylation occurred when the single Cys residue in S100A8 was mutated to Ala. S100A8-SNO in human neutrophils treated with NO donors was confirmed by the biotin switch assay. The stable adduct transnitrosylated hemoglobin, indicating a role in NO transport. S100A8-SNO suppressed mast cell activation by compound 48/80; intravital microscopy was used to demonstrate suppression of leukocyte adhesion and extravasation triggered by compound 48/80 in the rat mesenteric microcirculation. Although S100A8 is induced in macrophages by LPS or IFN-gamma, the combination, which activates inducible NO synthase, did not induce S100A8. Thus, the antimicrobial functions of NO generated under these circumstances would not be compromised by S100A8. Our results suggest that S100A8-SNO may regulate leukocyte-endothelial cell interactions in the microcirculation, and suppression of mast cell-mediated inflammation represents an additional anti-inflammatory property for S100A8. %Z FOR Codes: 111502 %0 Journal Article %~ PubMed %A Kalinowski, Danuta S %A Sharpe, Philip C %A Bernhardt, Paul V %A Richardson, Des R %T Structure-activity relationships of novel iron chelators for the treatment of iron overload disease: the methyl pyrazinylketone isonicotinoyl hydrazone series. %B Journal of Medicinal Chemistry %D 2008 %C United States %I Amer Chemical Soc %V 51 %N 2 %P 331-344 %@ 0022-2623 %X The design of novel Fe chelators with high Fe mobilization efficacy and low toxicity remains an important priority for the treatment of Fe overload disease. We have designed and synthesized the novel methyl pyrazinylketone isonicotinoyl hydrazone (HMPIH) analogs based on previously investigated aroylhydrazone chelators. The HMPIH series demonstrated high Fe mobilization efficacy from cells and showed limited to moderate antiproliferative activity. Importantly, this novel series demonstrated irreversible electrochemistry, which was attributed to the electron-withdrawing effects of the noncoordinating pyrazine N-atom. The latter functionality played a major role in forming redox-inactive complexes that prevent reactive oxygen species generation. In fact, the Fe complexes of the HMPIH series prevented the oxidation of ascorbate and hydroxylation of benzoate. We determined that the incorporation of electron-withdrawing groups is an important feature in the design of N, N, O-aroylhydrazones as candidate drugs for the treatment of Fe overload disease. %Z FOR Codes: 110199 %0 Journal Article %~ PubMed %A Whitnall, Megan %A Rahmanto, Yohan Suryo %A Sutak, Robert %A Xu, Xiangcong %A Becker, Erika M %A Mikhael, Marc R %A Ponka, Prem %A Richardson, Des R %T The MCK mouse heart model of Friedreich's ataxia: Alterations in iron-regulated proteins and cardiac hypertrophy are limited by iron chelation. %B Proceedings of the National Academy of Sciences of the United States of America %D 2008 %C US %I National Academy of Sciences %V 105 %N 28 %P 9757-62 %@ 1091-6490 %X There is no effective treatment for the cardiomyopathy of the most common autosomal recessive ataxia, Friedreich''s ataxia (FA). The identification of potentially toxic mitochondrial (MIT) iron (Fe) deposits in FA suggests that Fe plays a role in its pathogenesis. This study used the muscle creatine kinase conditional frataxin (Fxn) knockout (mutant) mouse model that reproduces the classical traits associated with cardiomyopathy in FA. We examined the mechanisms responsible for the increased cardiac MIT Fe loading in mutants. Moreover, we explored the effect of Fe chelation on the pathogenesis of the cardiomyopathy. Our investigation showed that increased MIT Fe in the myocardium of mutants was due to marked transferrin Fe uptake, which was the result of enhanced transferrin receptor 1 expression. In contrast to the mitochondrion, cytosolic ferritin expression and the proportion of cytosolic Fe were decreased in mutant mice, indicating cytosolic Fe deprivation and markedly increased MIT Fe targeting. These studies demonstrated that loss of Fxn alters cardiac Fe metabolism due to pronounced changes in Fe trafficking away from the cytosol to the mitochondrion. Further work showed that combining the MIT-permeable ligand pyridoxal isonicotinoyl hydrazone with the hydrophilic chelator desferrioxamine prevented cardiac Fe loading and limited cardiac hypertrophy in mutants but did not lead to overt cardiac Fe depletion or toxicity. Fe chelation did not prevent decreased succinate dehydrogenase expression in the mutants or loss of cardiac function. In summary, we show that loss of Fxn markedly alters cellular Fe trafficking and that Fe chelation limits myocardial hypertrophy in the mutant. %Z FOR Codes: 110106 %0 Book %A Rahmanto, Yohan %A Richardson, Des %T The Physiological and Pathophysiological Roles of Melanotransferrin %B %D 2008 %C Germany %I VDM Verlag %V %N %P %@ 9783639046007 %X %Z FOR Codes: 111699 110199 %0 Book Section %A Rahmanto, Yohan %A Sekyere, E O %A Dunne, L L %A Richardson, Des %T The function of the membrane-bound transferrin homologue, melanotransferin (melanoma tumour antigen p97) %B Iron Metabolism and Disease %D 2008 %C United States %I Transworld Research %V %N %P 219-239 %@ 9788130800660 %E Fuchs, H %X %Z FOR Codes: 110199 %0 Journal Article %~ PubMed %A Kovacevic, Zaklina %A Fu, Dong %A Richardson, Des R %T The iron-regulated metastasis suppressor, Ndrg-1: Identification of novel molecular targets. %B Biochimica et biophysica acta %D 2008 %C Netherlands %I Elsevier BV %V 1783 %N 10 %P 1981-92 %@ 0006-3002 %X A recently identified metastasis suppressor, N-myc downstream regulated gene-1 (Ndrg-1), has been shown to reduce the invasion and metastasis of breast, colon, prostate and pancreatic cancer. Among its many functions, Ndrg-1 is involved in modulating differentiation, proliferation and angiogenesis. However, knowledge of the molecular targets of Ndrg-1 is limited. The current study has focused on examining the functions of Ndrg-1 in a number of different cancer cell models including prostate, colon, lung and pancreatic cancer to elucidate the known pleiotropic nature of this protein. Furthermore, the potential gene targets of Ndrg-1 were analyzed using whole genome gene array revealing a substantial number of genes whose expression was affected by this metastasis suppressor. Significantly, Ndrg-1 up-regulated thiamine triphosphatase (Thtpa) expression in three of the four cell models. Thtpa is known to decrease the levels of the energy currency molecule, thiamine triphosphate, suggesting a potential pathway for the anti-proliferative effects of Ndrg-1. Furthermore, Ndrg-1 reduced the protein levels of cathepsin C which plays a role in invasion, indicating a potential mechanism of its anti-metastatic role in pancreatic cancer cells. These findings provide a potential link between the observed functions of Ndrg-1 and its molecular targets, further demonstrating its anti-metastatic effect. %Z FOR Codes: 110316 %0 Journal Article %~ PubMed %A Richardson, Des R %A Lok, Hiu Chuen %T The nitric oxide-iron interplay in mammalian cells: transport and storage of dinitrosyl iron complexes. %B Biochimica et Biophysica Acta %D 2008 %C Netherlands %I Elsevier BV %V 1780 %N 4 %P 638-651 %@ 0006-3002 %X Nitrogen monoxide (NO) is a vital effector and messenger molecule that plays roles in a variety of biological processes. Many of the functions of NO are mediated by its high affinity for iron (Fe) in the active centres of proteins. Indeed, NO possesses a rich coordination chemistry with this metal and the formation of dinitrosyl-dithiolato-Fe complexes (DNICs) is well known to occur intracellularly. In mammals, NO produced by activated macrophages acts as a cytotoxic effector against tumour cells by binding and releasing cancer cell Fe that is vital for proliferation. Glucose metabolism and the subsequent generation of glutathione (GSH) are critical for NO-mediated Fe efflux and this process occurs by active transport. Our previous studies showed that GSH is required for Fe mobilisation from tumour cells and we hypothesized it was effluxed with Fe as a dinitrosyl-diglutathionyl-Fe complex (DNDGIC). It is well known that Fe and GSH release from cells induces apoptosis, a crucial property for a cytotoxic effector like NO. Furthermore, NO-mediated Fe release is mediated from cells expressing the GSH transporter, multi-drug resistance protein 1 (MRP1). Interestingly, the glutathione-S-transferase (GST) enzymes act to bind DNDGICs with high affinity and some members of the GST family act as storage intermediates for these complexes. Since the GST enzymes and MRP1 form a coordinated system for removing toxic substances from cells, it is possible to hypothesize these molecules regulate NO levels by binding and transporting DNDGICs. %Z FOR Codes: 110199 %0 Journal Article %~ PubMed %A Bernhardt, Paul V %A Wilson, Gregory J %A Sharpe, Philip C %A Kalinowski, Danuta S %A Richardson, Des R %T Tuning the antiproliferative activity of biologically active iron chelators: characterization of the coordination chemistry and biological efficacy of 2-acetylpyridine and 2-benzoylpyridine hydrazone ligands. %B Journal of biological inorganic chemistry : JBIC : a publication of the Society of Biological Inorganic Chemistry %D 2008 %C Germany %I Springer %V 13 %N 1 %P 107-19 %@ 0949-8257 %X 2-Pyridinecarbaldehyde isonicotinoyl hydrazone (HPCIH) and di-2-pyridylketone isonicotinoyl hydrazone (HPKIH) are two Fe chelators with contrasting biological behavior. HPCIH is a well-tolerated Fe chelator with limited antiproliferative activity that has potential applications in the treatment of Fe-overload disease. In contrast, the structurally related HPKIH ligand possesses significant antiproliferative activity against cancer cells. The current work has focused on understanding the mechanisms of the Fe mobilization and antiproliferative activity of these hydrazone chelators by synthesizing new analogs (based on 2-acetylpyridine and 2-benzoylpyridine) that resemble both series and examining their Fe coordination and redox chemistry. The Fe mobilization activity of these compounds is strongly dependent on the hydrophobicity and solution isomeric form of the hydrazone (E or Z). Also, the antiproliferative activity of the hydrazone ligands was shown to be influenced by the redox properties of the Fe complexes. This indicated that toxic Fenton-derived free radicals are important for the antiproliferative activity for some hydrazone chelators. In fact, we show that any substitution of the H atom present at the imine C atom of the parent HPCIH analogs leads to an increase in antiproliferative efficacy owing to an increase in redox activity. These substituents may deactivate the imine R-C=N-Fe (R is Me, Ph, pyridyl) bond relative to when a H atom is present at this position preventing nucleophilic attack of hydroxide anion, leading to a reversible redox couple. This investigation describes novel structure-activity relationships of aroylhydrazone chelators that will be useful in designing new ligands or fine-tuning the activity of others. %Z FOR Codes: 30299 %0 Journal Article %~ PubMed %A Kalinowski, Danuta %A Sharpe, Philip %A Bernhardt, Paul %A Richardson, Des %T Design, Synthesis, and Characterization of New Iron Chelators with Anti-Proliferative Activity: Structure-Activity Relationships of Novel Thiohydrazone Analogues. %B Journal of medicinal chemistry %D 2007 %C United States %I Amer Chemical Soc %V 50 %N 24 %P 6212-25 %@ 0022-2623 %X Di-2-pyridylketone isonicotinoyl hydrazone Fe chelators utilize the N,N,O-donor set and have moderate anti-proliferative effects. Their closely related N,N,S-thiosemicarbazone analogues, namely, the di-2-pyridylketone thiosemicarbazones, exhibit markedly increased anti-proliferative and redox activity, and this was thought to be due to the inclusion of a sulfur donor atom (Richardson, D. R. et al. J. Med. Chem. 2006, 49, 6510-6521). To further examine the effect of donor atom identity on anti-proliferative activity, we synthesized thiohydrazone analogues of extensively examined aroylhydrazone chelators. The O,N,S-thiohydrazones exhibited decreased anti-proliferative effects compared to their parent aroylhydrazones and reduced redox activity. In contrast, the N,N,S-thiohydrazones showed vastly increased anti-proliferative activity compared to their hydrazone analogues, being comparable to potent thiosemicarbazones. Additionally, N,N,S-thiohydrazone complexes had reversible FeIII/II couples and exhibited increased redox activity. These observations demonstrate that the N,N,S-donor set is critical for potent anti-proliferative efficacy. %Z FOR Codes: %0 Journal Article %~ PubMed %A Kalinowski, Danuta %A Yu, Yu %A Sharpe, Philip %A Islam, Mohammad %A Liao, Yi-Tyng %A Lovejoy, David %A Kumar, Naresh %A Bernhardt, Paul %A Richardson, Des %T Design, Synthesis, and Characterization of Novel Iron Chelators: Structure-Activity Relationships of the 2-Benzoylpyridine Thiosemicarbazone Series and Their 3-Nitrobenzoyl Analogues as Potent Antitumor Agents. %B Journal of medicinal chemistry %D 2007 %C United States %I Amer Chemical Soc %V 50 %N 15 %P 3716-29 %@ 0022-2623 %X Previously, we demonstrated that the potent antiproliferative activity of the di-2-pyridylketone thiosemicarbazone (DpT) series of Fe chelators was due to their ability to induce Fe depletion and form redox-active Fe complexes (Richardson, D. R.; et al. J. Med. Chem. 2006, 49, 6510-6521). We now examine the role of aromatic substituents on the antiproliferative and redox activity of novel DpT analogues, namely, the 2-benzoylpyridine thiosemicarbazone (BpT) and 2-(3-nitrobenzoyl)pyridine thiosemicarbazone (NBpT) series. Both series exhibited selective antiproliferative effects, with the majority having greater antineoplastic activity than their DpT homologues. This makes the BpT chelators the most active anticancer agents developed within our laboratory. The BpT series Fe complexes exhibit lower redox potentials than their corresponding DpT and NBpT complexes, highlighting their enhanced redox activity. The increased ability of BpT-Fe complexes to catalyze ascorbate oxidation and benzoate hydroxylation, relative to their DpT and NBpT analogues, suggested that redox cycling plays an important role in their antiproliferative activity. %Z FOR Codes: %0 Journal Article %~ PubMed %A Mrkvickov, Zlata %A Kovarkov, Petra %A Klimes, Jir %A Kalinowski, Danuta %A Richardson, Des R %T Development and validation of HPLC-DAD methods for the analysis of two novel iron chelators with potent anti-cancer activity. %B Journal of pharmaceutical and biomedical analysis %D 2007 %C Netherlands %I Elsevier BV %V 43 %N 4 %P 1343-1351 %@ 0731-7085 %X Di-2-pyridylketone isonicotinoyl hydrazone (PKIH) and di-2-pyridylketone-4,4-dimethyl-3-thiosemicarbazone (Dp44mT) novel iron chelators which possess marked anti-cancer activity in vivo. However, further progress in the development of these drug candidates requires precise and convenient methods for their qualitative and quantitative analysis. The aim of this study was to develop and validate HPLC methods suitable for the purity and stability evaluation of Dp44mT and PKIH and subsequently to employ these methods in stress tests addressing their chemical stability. The chromatographic analyses of both chelators were accomplished via HPLC using a Discovery HSF5 column (25cmx4mm; 5mum). For separation of Dp44mT and its synthetic precursors, the mobile phase was composed of a mixture of 2mM EDTA and acetonitrile in a ratio 60:40 (v/v). A desirable separation of PKIH from its synthetic precursors was achieved with a mixture of 0.01M phosphate buffer (pH 3.0), methanol and acetonitrile in a ratio of 65:21:14 (v/v/v) with the addition of EDTA (2mM). In order to confirm the utility of these HPLC methods for measuring these drugs and their stability, Dp44mT and PKIH were subjected to chemical stress tests. These experiments showed that Dp44mT was relatively stable against hydrolytic degradation, but quite sensitive to oxidation. On the other hand, PKIH was slightly sensitive to acid-catalyzed hydrolysis, but it was relatively stable under other tested conditions. Furthermore, these studies confirmed the utility of these methods not only for appropriate evaluation of purity but also stability. The analytical methods developed and validated in this study, as well as the basic data on the chemical stability, should further support the development of both these novel anti-cancer chelators as promising drug candidates. %Z FOR Codes: %0 Journal Article %~ PubMed %A Richardson, Des R %A Suryo Rahmanto, Yohan %T Differential regulation of the Menkes and Wilson disease copper transporters by hormones: an integrated model of metal transport in the placenta. %B Biochemical Journal %D 2007 %C United Kingdom %I Portland Press Ltd. %V 402 %N 2 %P e1-e3 %@ 1470-8728 %X Copper (Cu) plays a critical role in the developing foetus, but virtually nothing is known concerning the regulation of its uptake and metabolism in the placenta. In this issue of the Biochemical Journal, Hardman and colleagues, using a model of placental trophoblasts in culture, identify differential hormonal regulation of two copper-transporting ATPases; namely, those responsible for Menkes disease (ATP7A; MNK) and Wilson disease (ATP7B; WND). Insulin and oestrogen, which are essential during gestation, up-regulate MNK and this leads to trafficking of the MNK protein from the Golgi to the basolateral membrane, resulting in increased Cu efflux. At the same time, insulin decreased WND levels, and this leads to intracellular sequestration of the protein to a perinuclear region that reduces apical Cu release. As such, this results in a concerted flux of Cu from the basolateral surface of the trophoblast that would potentially be used by the developing foetus. An integrated model of vectorized Cu transport is proposed, which involves co-ordinated expression of transporters, organelle interactions and probable protein-protein interactions. The findings have wider implications for considering general models of intracellular metal transport. %Z FOR Codes: 60108 %0 Journal Article %~ PubMed %A Kalinowski, Danuta S %A Richardson, Des R %T Future of toxicology--iron chelators and differing modes of action and toxicity: the changing face of iron chelation therapy. %B Chemical research in toxicology %D 2007 %C United States %I American Chemical Society %V 20 %N 5 %P 715-20 %@ 0893-228X %X Iron (Fe) chelation therapy was initially designed to alleviate the toxic effects of excess Fe evident in Fe-overload diseases. However, the novel toxicological properties of some Fe chelator-metal complexes have shifted appreciable focus to their application in cancer chemotherapy. Redox-inactive Fe chelator complexes are well suited for the treatment of Fe-overload diseases, whereas Fe chelator complexes with high redox activity have shown promising results as chemotherapeutics against cancer. Within this perspective, we discuss the different modes of action and toxicological profiles of Fe chelators, including analogues of 2-pyridylcarboxaldehyde isonicotinoyl hydrazone, di-2-pyridylketone isonicotinoyl hydrazone, di-2-pyridylketone thiosemicarbazone, and the clinically trialed chelator 3-aminopyridine-2-carboxaldehyde thiosemicarbazone. The potential application of these agents in the changing face of Fe chelation therapy is discussed. %Z FOR Codes: %0 Journal Article %~ Isi %A Sharpe, PC %A Richardson, DR %A Bernhardt, PV %A Chin, P %T Hydrazone chelators for the treatment of iron overload disorders: iron coordination chemistry and biological activity %B DALTON TRANSACTIONS %D 2007 %C United Kingdom %I Royal Society of Chemistry %V 30 %N %P 3232-3244 %@ 1477-9226 %X %Z FOR Codes: %0 Journal Article %~ PubMed %A Suryo Rahmanto, Y %A Dunn, Ll %A Richardson, Dr %T Identification of Distinct Changes in Gene Expression after Modulation of Melanoma Tumor Antigen p97 (Melanotransferrin) in Multiple Models In Vitro and In Vivo. %B Carcinogenesis %D 2007 %C United States %I Lippincott Williams and Wilkins %V 28 %N 10 %P 2172-83 %@ 0143-3334 %X Melanoma tumor antigen p97 or melanotransferrin (MTf) is an iron (Fe)-binding protein with high homology to serum transferrin. MTf is expressed at very low levels in normal tissues and in high amounts in melanoma cells although its function remains elusive. To understand the function of MTf, we utilized whole-genome microarray analysis to examine the gene expression profile of five models after modulating MTf expression. These models included two new stably transfected MTf hyper-expression models (SK-N-MC neuroepithelioma and LMTK- fibroblasts) and one cell type (SK-Mel-28 melanoma) where MTf was down-regulated by post-transcriptional gene silencing. These findings were compared with alterations in gene expression identified using the MTf-/- mice. In addition, the changes identified from the microarray data were also assessed in a new model of MTf down-regulation in SK-Mel-2 melanoma cells. In the cell line models, MTf hyper-expression led to increased proliferation, whereas MTf down-regulation resulted in decreased proliferation. Across all five models of MTf down- and up-regulation, we identified three genes modulated by MTf. These included ATP-binding cassette subfamily B member 5, whose change in expression mirrored MTf down- or up-regulation. In addition, thiamine triphosphatase and transcription factor 4 were inversely expressed relative to MTf levels across all five models. The products of these three genes are involved in membrane transport, thiamine phosphorylation and proliferation/survival, respectively. This study identifies novel molecular targets directly or indirectly regulated by MTf and the potential pathways involved in its function, including modulation of proliferation. %Z FOR Codes: 111201 %0 Journal Article %~ PubMed %A Musiol, Robert %A Jampilek, Josef %A Kralova, Katarina %A Richardson, Des R %A Kalinowski, Danuta %A Podeszwa, Barbara %A Finster, Jacek %A Niedbala, Halina %A Palka, Anna %A Polanski, Jaroslaw %T Investigating biological activity spectrum for novel quinoline analogues. %B Bioorganic & medicinal chemistry %D 2007 %C United Kingdom %I Pergamon-Elsevier Science Ltd %V 15 %N 3 %P 1280-1288 %@ 0968-0896 %X The lack of the wide spectrum of biological data is an important obstacle preventing the efficient molecular design. Quinoline derivatives are known to exhibit a variety of biological effects. In the current publication, we tested a series of novel quinoline analogues for their photosynthesis-inhibiting activity (the inhibition of photosynthetic electron transport in spinach chloroplasts (Spinacia oleracea L.) and the reduction of chlorophyll content in Chlorella vulgaris Beij.). Moreover, antiproliferative activity was measured using SK-N-MC neuroepithelioma cell line. We described the structure-activity relationships (SAR) between the chemical structure and biological effects of the synthesized compounds. We also measured the lipophilicity of the novel compounds by means of the RP-HPLC and illustrate the relationships between the RP-HPLC retention parameter logK (the logarithm of capacity factor K) and logP data calculated by available programs. %Z FOR Codes: 110199 %0 Journal Article %~ PubMed %A Podeszwa, B %A Niedbala, H %A Polanski, J %A Musiol, R %A Tabak, D %A Finster, J %A Serafin, K %A Milczarek, M %A Wietrzyk, J %A Boryczka, S %A Mol, W %A Jampilek, J %A Dohnal, J %A Kalinowski, D S %A Richardson, D R %T Investigating the antiproliferative activity of quinoline-5,8-diones and styrylquinolinecarboxylic acids on tumor cell lines. %B Bioorganic & medicinal chemistry letters %D 2007 %C United Kingdom %I Pergamon %V 17 %N 22 %P 6138-6141 %@ 0960-894X %X The structure-activity relationships of new quinoline based compounds were investigated. Quinoline-5,8-dione and styrylquinoline scaffolds were used for the design of potentially active compounds. The novel analogues had comparable antiproliferative activity to cisplatin when evaluated in a bioassay against the P388 leukemia cell line. However, these compounds appeared far less efficient against SK-N-MC neuroepithelioma cells. Analogues without the 5,8-dione structure but containing the 8-carboxylic acid group were also found to induce antiproliferative activity. Hydrophobicity as measured by HPLC did not correlate with antiproliferative activity. %Z FOR Codes: 110199 %0 Journal Article %~ PubMed %A Xu, Xiangcong %A Sutak, Robert %A Richardson, Des R %T Iron Chelation by Clinically Relevant Anthracyclines: Alteration in Expression of Iron-Regulated Genes and Atypical Changes in Intracellular Iron Distribution and Trafficking. %B Molecular pharmacology %D 2007 %C United States %I Amer Soc Pharmacology Experimental Therapeutics %V 73 %N 3 %P 833-44 %@ 0026-895X %X Anthracyclines are effective anticancer agents. However, their use is limited by cardiotoxicity, an effect linked to their ability to chelate iron and to perturb iron metabolism (Mol Pharmacol 68:261-271, 2005). These effects on iron-trafficking remain poorly understood, but they are important to decipher because treatment for anthracycline cardiotoxicity uses the chelator, dexrazoxane. Incubation of cells with doxorubicin (DOX) up-regulated mRNA levels of the iron-regulated genes transferrin receptor-1 (TfR1) and N-myc downstream-regulated gene-1 (Ndrg1). This effect was mediated by iron depletion, because it was reversed by adding iron and it was prevented by saturating the anthracycline metal binding site with iron. However, DOX did not act like a typical chelator, because it did not induce cellular iron mobilization. In the presence of DOX and (59)Fe-transferrin, iron-trafficking studies demonstrated ferritin-(59)Fe accumulation and decreased cytosolic-(59)Fe incorporation. This could induce cytosolic iron deficiency and increase TfR1 and Ndrg1 mRNA. Up-regulation of TfR1 and Ndrg1 by DOX was independent of anthracycline-mediated radical generation and occurred via hypoxia-inducible factor-1alpha-independent mechanisms. Despite increased TfR1 and Ndrg1 mRNA after DOX treatment, this agent decreased TfR1 and Ndrg1 protein expression. Hence, the effects of DOX on iron metabolism were complex because of its multiple effector mechanisms. %Z FOR Codes: %0 Journal Article %~ PubMed %A Fu, Dong %A Richardson, Des R %T Iron chelation and regulation of the cell cycle: 2 mechanisms of posttranscriptional regulation of the universal cyclin-dependent kinase inhibitor p21CIP1/WAF1 by iron depletion. %B Blood %D 2007 %C United States %I American Society of Hematology %V 110 %N 2 %P 752-761 %@ 1528-0020 %X Iron (Fe) plays a critical role in proliferation, and Fe deficiency results in G(1)/S arrest and apoptosis. However, the precise role of Fe in cell-cycle control remains unclear. We observed that Fe depletion increased the mRNA of the universal cyclin-dependent kinase inhibitor, p21(CIP1/WAF1), while its protein level was not elevated. This observation is unique to the G(1)/S arrest seen after Fe deprivation, as increased p21(CIP1/WAF1) mRNA and protein are usually found when arrest is induced by other stimuli. In this study, we examined the posttranscriptional regulation of p21(CIP1/WAF1) after Fe depletion and demonstrated that its down-regulation was due to 2 mechanisms: (1) inhibited translocation of p21(CIP1/WAF1) mRNA from the nucleus to cytosolic translational machinery; and (2) induction of ubiquitin-independent proteasomal degradation. Iron chelation significantly (P < .01) decreased p21(CIP1/WAF1) protein half-life from 61 (+/- 4 minutes; n = 3) to 28 (+/- 9 minutes, n = 3). Proteasomal inhibitors rescued the chelator-mediated decrease in p21(CIP1/WAF1) protein, while lysosomotropic agents were not effective. In Fe-replete cells, p21(CIP1/WAF1) was degraded in an ubiquitin-dependent manner, while after Fe depletion, ubiquitin-independent proteasomal degradation occurred. These results are important for considering the mechanism of Fe depletion-mediated cell-cycle arrest and apoptosis and the efficacy of chelators as antitumor agents. %Z FOR Codes: %0 Journal Article %~ Isi %A Jerebtsova, M %A Richardson, DR %A Nekhai, S %A Gordeuk, VR %A Ray, PE %A Debebe, Z %A Kurantsin-Mills, J %A Ammosova, T %A Charles, S %A Niu, XM %T Iron chelators ICL670 and 311 inhibit HIV-1 transcription %B VIROLOGY %D 2007 %C United States %I Academic Press %V 367 %N 2 %P 324-333 %@ 0042-6822 %X %Z FOR Codes: %0 Journal Article %~ PubMed %A Suryo Rahmanto, Y %A Dunn, L L %A Richardson, D R %T The melanoma tumor antigen, melanotransferrin (p97): a 25-year hallmark - from iron metabolism to tumorigenesis. %B Oncogene %D 2007 %C United Kingdom %I Nature Publishing Group %V 26 %N 42 %P 6113-24 %@ 1476-5594 %X Melanotransferrin (MTf) or melanoma tumor antigen p97 is a transferrin (Tf) homolog that is found predominantly bound to the cell membrane via a glycosyl phosphatidylinositol anchor. The molecule is a member of the Tf superfamily and binds iron through a single high-affinity iron(III)-binding site. Since its discovery on the plasma membrane of melanoma cells, the function of MTf has remained intriguing, particularly in relation to its role in cancer cell iron transport. In fact, considering the crucial role of iron in many metabolic pathways, e.g., DNA synthesis, it was important to understand the function of MTf in the transport of this vital nutrient. MTf has also been implicated in diverse physiological processes, such as plasminogen activation, angiogenesis and cell migration. However, recent studies using a knockout mouse and post-transcriptional gene silencing have demonstrated that MTf is not involved in iron metabolism, but plays a vital role in melanoma cell proliferation and tumorigenesis. In this review, we discuss the possible biological functions of MTf, particularly in relation to cancer. %Z FOR Codes: %0 Journal Article %~ PubMed %A Yu, Yu %A Kovacevic, Zaklina %A Richardson, Des %T Tuning Cell Cycle Regulation With an Iron Key. %B Cell cycle (Georgetown, Tex.) %D 2007 %C United States %I Landes Bioscience %V 6 %N 16 %P 1982-94 %@ 1551-4005 %X Iron (Fe) is essential for cellular metabolism e.g., DNA synthesis and its depletion causes G(1)/S arrest and apoptosis. Considering this, Fe chelators have been shown to be effective anti-proliferative agents. In order to understand the anti-tumor activity of Fe chelators, the mechanisms responsible for G(1)/S arrest and apoptosis after Fe-depletion have been investigated. These studies reveal a multitude of cell cycle control molecules are regulated by Fe. These include p53, p27(Kip1), cyclin D1 and cyclin-dependent kinase 2(cdk2). Additionally, Fe-depletion up-regulates the mRNA levels of the cdk inhibitor, p21(CIP1/WAF1), but paradoxically down-regulates its protein expression. This effect could contribute to the apoptosis observed after Fe-depletion. Iron-depletion also leads to proteasomal degradation of p21(CIP1/WAF1) and cyclin D1 via an ubiquitin-independent pathway. This is in contrast to the mechanism in Fe-replete cells, where it occurs by ubiquitin-dependent proteasomal degradation. Up-regulation of p38 mitogen-activated protein kinase (MAPK) after Fe-depletion suggests another facet of cell cycle regulation responsible for inhibition of proliferation and apoptosis induction. Elucidation of the complex effects of Fe-depletion on the expression of cell cycle control molecules remains at its infancy. However, these processes are important to dissect for complete understanding of Fe-deficiency and the development of chelators for cancer treatment. %Z FOR Codes: