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Mestrado Bolonha em Gestão e Estratégia Industrial Recentemente, vários estudos começaram a avaliar empiricamente a agilidade, adaptabilidade e o alinhamento das cadeias de abastecimento. Com a pandemia Covid19 as cadeias de abastecimento que eram consideradas ágeis, adaptáveis e alinháveis apresentaram um resultando na melhoria do desempenho de interrupção pós-COVID. O que destacou que a capacidade de simultaneamente ser ágil, adaptável e alinhar toda a cadeia de abastecimento para desenvolver uma vantagem competitiva sustentável. Sendo assim, este trabalho tem como principal objetivo estudar o impacto da digitalização nas caraterísticas da adaptabilidade, agilidade, o alinhamento, flexibilidade, inovação das cadeias de abastecimento e consequentemente no seu desempenho na mesma. Primeiramente, os resultados obtidos permitiram concluir que a orientação digital e a digitalização têm um impacto positivo com a agilidade, adaptação, alinhamento, flexibilidade e a inovação da cadeia de abastecimento. Posteriormente, foi ainda possível verificar que a agilidade e a adaptação não possuem um impacto significativo no desempenho da cadeia de abastecimento. Este resultado contrariou diversos estudos que defendiam que a agilidade e a adaptação têm de facto uma relação positiva com o desempenho da cadeia de abastecimento, quer seja por um custo eficiente ou pelo serviço ao cliente. Por sua vez, obteve-se como resultado que o alinhamento, a flexibilidade e a inovação da cadeia de abastecimento detêm de um impacto significativo no desempenho da cadeia de abastecimento, enaltecendo assim diversos estudos anteriores. Recently, several studies have begun to empirically evaluate the agility, adaptability, and alignment of supply chains. With the Covid-19 pandemic the supply chains that were found to be agile, adaptable and alignable showed a resultant improvement in disruption performance post-COVID. Which highlighted that the ability to simultaneously be agile, adaptive and coordinate the entire supply chain to develop a sustainable competitive advantage. Therefore, the main objective of this paper is to study the effect of digitalization on various characteristics such as flexibility, innovation, adaptability, agility and alignment of supply chains and consequently on their performance. Firstly, the results obtained allowed us to conclude that digital orientation and digitalization have a positive impact on agility, adaptation, alignment, flexibility and innovation of the supply chain. Subsequently, it was further found that Agility and Adaptation do not have a significant impact on supply chain performance. This result contradicted several studies which argued that agility and adaptation have indeed a positive relationship with supply chain performance, either by cost efficiency or by customer service. In turn, it was obtained as a result that supply chain alignment, flexibility and innovation have a significant impact on supply chain performance, thus extolling several previous studies. info:eu-repo/semantics/publishedVersion

Gramibactin (GBT) is an archetype for the new class of diazeniumdiolate siderophores, produced by Paraburkholderia graminis, a cereal‐associated rhizosphere bacterium, for which a detailed solution thermodynamic study exploring the iron coordination properties is reported. The acid‐base behavior of gramibactin as well as its complexing ability toward Fe3+ was studied over a wide range of pH values (2≤pH≤11). For the latter the ligand‐competition method employing EDTA was used. Only two species are formed: [Fe(GBT)]− (pH 2 to 9) and [Fe(GBT)(OH)2]3− (pH≥9). The formation of [Fe(GBT)]− and its occurrence in real systems was confirmed by LC‐HRESIMS analysis of the bacteria culture broth extract. The sequestering ability of gramibactin was also evaluated in terms of the parameters pFe and pL0.5. Gramibactin exhibits a higher sequestering ability toward Fe3+ than EDTA and of the same order of magnitude as hydroxamate‐type microbial siderophores, but smaller than most of the catecholate‐type siderophores and much higher than the most known phytosiderophores., The sequestration properties of gramibactin as a prototype for the new class of diazeniumdiolate siderophores is presented and compared with siderophores derived from catecholates and hydroxamates. According to pFe and pL0.5 values, gramibactin is a considerably more potent sequestering agent than the known class of phytosiderophores and approximately reaches the ability of typical hydroxamates like desferrioxamines.

Organic matter regulates the availability of important trace elements in aquatic and terrestrial ecosystems by acting as a source and container for microbes. To overcome the limitation of trace elements, nitrogen-fixing bacteria, e.g. release low-molecular-weight chelators (metallophores), which scavenge the essential cofactors of the nitrogenase, iron, and molybdenum (Mo), via complexation and subsequent uptake. The formation of metallophores is triggered by limiting conditions, which must be replicated in the laboratory in order to study metallophores as a mediator in metal cycling. While ethylenediaminetetraacetic acid (EDTA)-based buffer systems for metal cations are well established, there is limited knowledge regarding the buffering of oxoanions such as molybdate in a bacterial growth medium. To mimic the availability of molybdenum in nature under laboratory conditions, this study created a Mo-buffer system for bacterial growth media of the model organisms Azotobacter vinelandii and Frankia sp. CH37. We investigated selected hydroxypyridinones (HPs) as potential molybdenum-chelating agents, determining the amount required for efficient molybdenum complexation by calculating speciation plots of the various candidate complexes in artificial growth media at various pH values. The Mo–maltol system was identified as an ideal, nontoxic molybdenum-buffer system. In the presence of the Mo–maltol system, the growth of Frankia sp. was limited under diazotrophic conditions, whereas A. vinelandii could acquire molybdenum through the release of protochelin and subsequent molybdenum uptake. The study paves the way for unravelling molybdenum recruitment and homeostasis under limiting conditions in bacteria.

Tese de mestrado, Medicina Dentaria, Universidade de Lisboa, Faculdade de Medicina Dentaria, 2020 Submitted by Luiza Baptista (luiza.baptista@fmd.ulisboa.pt) on 2021-03-01T15:35:08Z No. of bitstreams: 1 Dissertação_MIMD_Inês_Gama_Silva_2019-2020.pdf: 520097 bytes, checksum: 7086e2e2d2a634fdc85c334b95639c86 (MD5) Made available in DSpace on 2021-03-01T15:35:34Z (GMT). No. of bitstreams: 1 Dissertação_MIMD_Inês_Gama_Silva_2019-2020.pdf: 520097 bytes, checksum: 7086e2e2d2a634fdc85c334b95639c86 (MD5) Previous issue date: 2020-11-18

Soil pollution by metal ions constitutes one of the most significant environmental problems in the world, being the ecosystems of extended areas wholly compromised. The remediation of soils is an impelling necessity, and different methodologies are used and studied for reaching this goal. Among them, the application of chelating agents is one of the most promising since it could allow the removal of metal ions while preserving the most meaningful properties of the original soils. The research in this field requires the joined contribute of different expertise spanning from biology to chemistry. In this work, we propose a parsimonious and pragmatic approach for screening among a range of potential chelating agents. This methodology, the Nurchi's method, is based on an extension of the Reilley procedure for EDTA titrations. This allows forecasting the binding ability of chelating agents toward the target polluting metal ions and those typically found in soils, based on the knowledge of the related protonation and complex formation constants. The method is thoroughly developed, and then tested by application to some representative cases. Its use and relevance in biomedical and industrial applications is also discussed.

The interpretation of in vitro cytotoxicity data of Cu(II)-1,10-phenanthroline (phen) complexes normally does not take into account the speciation that complexes undergo in cell incubation media and its implications in cellular uptake and mechanisms of action. We synthesize and test the activity of several distinct Cu(II)-phen compounds; up to 24 h of incubation, the cytotoxic activity differs for the Cu complexes and the corresponding free ligands, but for longer incubation times (e.g., 72 h), all compounds display similar activity. Combining the use of several spectroscopic, spectrometric, and electrochemical techniques, the speciation of Cu-phen compounds in cell incubation media is evaluated, indicating that the originally added complex almost totally decomposed and that Cu(II) and phen are mainly bound to bovine serum albumin. Several methods are used to disclose relationships between structure, activity, speciation in incubation media, cellular uptake, distribution of Cu in cells, and cytotoxicity. Contrary to what is reported in most studies, we conclude that interaction with cell components and cell death involves the separate action of Cu ions and phen molecules, not [Cu(phen)(n)] species. This conclusion should similarly apply to many other Cu-ligand systems reported to date. Fundacao para a Ciencia e Tecnologia (FCT)Portuguese Foundation for Science and TechnologyEuropean Commission [UID/Multi/04349/2019, UIDB/00100/2020, UID/QUI/50006/2019, UID/BIO/04565/2020] FCTPortuguese Foundation for Science and TechnologyEuropean Commission [IF/00841/2012, IF/00007/2015, SFRH/BD/108743/2015] Programa Operacional Regional de Lisboa 2020 [007317] PROTEOMASS Scientific Society COST Action, NECTAR (Network for Equilibria and Chemical Thermodynamics Advanced Research) - COST (European Cooperation in Science and Technology) [CA18202] info:eu-repo/semantics/publishedVersion

8-hydroxyquinoline-2-carboxylic acid (8-HQA) has been found in high concentrations (0.5&ndash, 5.0 mmol&middot, dm&minus, 3) in the gut of Noctuid larvae (and in a few other lepidopterans), in which it is proposed to act as a siderophore. Since it is known that many natural siderophores are also involved in the uptake and metabolism of other essential elements than iron, this study reports some results on the investigation of 8-HQA interactions with molybdate (MoO42&minus, i.e., the main molybdenum form in aqueous environments), in order to understand the possible role of this ligand as molybdophore. A multi-technique approach has been adopted, in order to derive a comprehensive set of information necessary to assess the chemical speciation of the 8-HQA/MoO42&minus, system, as well as the coordination behavior and the sequestering ability of 8-HQA towards molybdate. Chemical speciation studies have been performed in KCl(aq) at I = 0.2 mol&middot, 3 and T = 298.15 K by ISE-H+ (glass electrode) potentiometric and UV/Vis spectrophotometric titrations. CV (Cyclic Voltammetry), DP-ASV (Differential Pulse-Anodic Stripping Voltammetry), ESI-MS experiments and quantum mechanical calculations have been also performed to derive information about the nature and possible structure of species formed. These results are also compared with those reported for the 8-HQA/Fe3+ system in terms of chemical speciation and sequestering ability of 8-HQA.

The development of pharmacologically active compounds based on bis(thiosemicarbazones) (BTSC) and on their coordination to metal centers constitutes a promising field of research. We have recently explored this class of ligands and their Cu(II) complexes for the design of cancer theranostics agents with enhanced uptake by tumoral cells. In the present work, we expand our focus to aliphatic and aromatic BTSC Zn(II) complexes bearing piperidine/morpholine pendant arms. The new complexes ZnL1–ZnL4 were characterized by a variety of analytical techniques, which included single-crystal X-ray crystallography for ZnL2 and ZnL3. Taking advantage of the fluorescent properties of the aromatic complexes, we investigated their cellular uptake kinetics and subcellular localization. Furthermore, we tried to elucidate the mechanism of action of the cytotoxic effect observed in human cancer cell line models. The results show that the aliphatic complexes (ZnL1 and ZnL2) have a symmetrical structure, while the aromatic counterparts (ZnL3 and ZnL4) have an asymmetrical nature. The cytotoxic activity was higher for the aromatic BTSC complexes, as well as the cellular uptake, evaluated by measurement of intracellular Zn accumulation. Among the most active complexes, ZnL3 presented the fastest uptake kinetics and lysosomal localization assessed by live-cell microscopy. Detailed studies of its impact on cellular production of reactive oxygen species and impairment of lysosomal membrane integrity reinforced the influence of the pendant piperidine in the biological performance of aromatic BTSC Zn(II) complexes.

The recognition of quinolinic carboxylic acids as natural chelants and the recent observation of a high production of 8-hydroxyquinoline-2-carboxylic acid (8-HQA) in the gut of several Noctuid larvae (e.g. Spodoptera littoralis) has inspired the study of the chelation properties of 8-HQA towards Fe2+ and Fe3+. Here, we report a detailed characterization of the thermodynamic solution behaviour of Fe2+/8-HQA and Fe3+/8-HQA systems as a function of the pH value. The acid–base properties of 8-HQA and its binding ability towards Fe2+ and Fe3+ have been investigated over a wide range of pH values (2.0 ≤ pH ≤ 11.0) by ISE-H+ (glass electrode) potentiometric titrations in KCl(aq) at I = 0.2 mol dm−3 and at T = 298.15 K. For both oxidation states, various FeLqHr species are formed, with q = 1, 2 (and 3), and −2 ≤ r ≤ 1. The presence of the main FeLqHr species was confirmed by HESI-HRMS. ESR measurements have also been performed to get some extra information on the Fe3+ coordination, indicating a distorted octahedral symmetry around the metal center. Quantum mechanical calculations have been carried out in order to characterize the structural features of selected metal complexes. The complexing ability of 8-HQA is generally much higher for Fe3+ than Fe2+. Nevertheless, the sequestering ability of 8-HQA towards these two oxidation states of this metal ion, obtained by the calculation of several pL0.5 values, resulted in it being highly dependent on the pH value: (i) at relatively low pH values, it is higher for Fe3+ (pL0.5 = 6.3 at pH = 3.0) than for Fe2+ (pL0.5 = 3.1 at pH = 3.0); (ii) it is almost the same at pH = 8.1 (Fe3+: pL0.5 = 8.3; Fe2+: pL0.5 = 8.1); (iii) it is higher for Fe2+ at high pH values (pL0.5 = 8.9 for Fe2+ and pL0.5 = 6.2 for Fe3+ at pH = 10.0). The determination of the stability constants of the Fe2+/8-HQA and Fe3+/8-HQA complexes was also complemented by data obtained by the ligand-competition approach, using EDTA as a competing ligand over a wide range of cation and ligand concentrations and ratios. This also allowed a more thorough investigation of both the Fe2+/EDTA and Fe3+/EDTA systems, providing an accurate stability constant dataset for the Fep(EDTA)qHr species under the above-mentioned experimental conditions, which are commonly used in biological studies.

We present the combination of the clinically well-proven chemotherapeutic agent, Doxorubicin, and (99m)Tc, an Auger and internal conversion electron emitter, into a dual-action agent for therapy. Chemical conjugation of Doxorubicin to (99m)Tc afforded a construct which autonomously ferries a radioactive payload into the cell nucleus. At this site, damage is exerted by dose deposition from Auger radiation. The (99m)Tc-conjugate exhibited a dose-dependent inhibition of survival in a selected panel of cancer cells and an in vivo study in healthy mice evidenced a biodistribution which is comparable to that of the parent drug. The homologous Rhenium conjugate was found to effectively bind to DNA, inhibited human Topoisomerase II, and exhibited cytotoxicity in vitro. The collective in vitro and in vivo data demonstrate that the presented metallo-conjugates closely mimic native Doxorubicin.

The microwave-assisted synthesis of Pt(II) complexes containing several pyridines (i.e., pyridine L1 , 2-picoline L2 , 3-picoline L3 , 4-picoline L4 , 2,2′-bipyridine L5 ) is reported. For L1 – L5 , the reaction was successful in about 50% yield with all the ligands except L2 . The same method applied to 4,4′-bis(2-morpholinoethoxy)-2,2′-bipyridine ( L6 , a ligand showing interesting antiproliferative properties because of a high DNA affinity), was unsatisfactory. The corresponding complex cis -[PtCl 2 ( L6 )] was obtained only heating at reflux a mixture of [PtCl 2 (1,5-cyclooctadiene)] and L6 in acetonitrile for 24 h. Antiproliferative activity of [PtCl 2 ( L6 )] on four cancer cell lines (ovarian A2780 and its cisplatin-resistant variant A2780cisR, prostate PC3 and breast cancer MDA-MB-231) was compared with that of its ligand and the model complex [PtCl 2 ( L5 )]. These studies showed that [PtCl 2 ( L6 )] has just marginal activity towards the tested cells if compared with cisplatin.

Genome mining and chemical analyses revealed that rhizosphere bacteria (Paraburkholderia graminis) produce a new type of siderophore, gramibactin, a lipodepsipeptide that efficiently binds iron with a logβ value of 27.6. Complexation-induced proton NMR chemical shifts show that the unusual N-nitrosohydroxylamine (diazeniumdiolate) moieties participate in metal binding. Gramibactin biosynthesis genes are conserved in numerous plant-associated bacteria associated with rice, wheat, and maize, which may utilize iron from the complex.

A novel series of platinum(II) complexes bearing aliphatic amines and ligands with DNA-targeting properties was synthesized to achieve more potent and selective metallodrugs. We developed six new platinum-based drugs, which contain methylamine, 1a-c, and isopropylamine, 2a-c, both in the trans position to a selected targeting ligand: naphthalimide. The activity of the complexes has been evaluated in order to confirm the improvements from our proposed approach, and the complexes demonstrate better cytotoxic activity on cancer cell lines when compared with the ligands and, importantly, with cisplatin. Further studies were performed to assess their subcellular localization and binding mode to DNA.

Seeking self-activating chemical nucleases with potential applications as therapeutic agents, new ternary terpyridine–bipyridine–Cu(II) complexes carrying pendant cyclic amines were developed. After detailed characterization, the nuclease activity of the synthesized compounds was evaluated by using circular plasmid DNA as substrate and analyzing the products by agarose-gel electrophoresis. The new complexes present an impressive plasmid DNA cleaving ability, which triggers double-strand DNA breaks in the absence of any exogenous agents, via an oxidative mechanism. The binding affinity towards duplex DNA was determined using UV-Vis and fluorescence spectroscopic titrations. These studies showed that the tested complexes bind moderately (in the order of 104 M−1) to duplex DNA. The copper complexes displayed high cytotoxicity against ovarian carcinoma A2780 cells (4-fold cisplatin activity), surpassing the resistance on the cisplatin-resistant cell line (A2780cisR) with lower resistance factors. Cellular uptake studies showed that the ternary complexes were able to enter the cell with a significant localization in the cytoskeleton.

The chemical and biological features of two newly synthesized [PtCl2(L)(2-aminonaphthalene)] complexes (L is NH3 or 2-aminonaphthalene) were compared with those of two already reported enantiomeric complexes of formula [PtCl2(DABN)] [DABN is (R)-1,1'-binaphthyl-2,2'-diamine or (S)-1,1'-binaphthyl-2,2'-diamine]. Solution behavior, lipophilicity, cytotoxicity with regard to one colorectal (HCT116) and two ovarian (A2780 and A2780Cp8) human carcinoma cell lines, and in vitro DNA- and G-quadruplex-binding properties were evaluated. In particular, the cytotoxicity of [PtCl2(NH3)(2-aminonaphthalene)] was better than that of cisplatin for all cell lines, and rather resembled that of oxaliplatin. The solution behavior of the whole series of complexes and the absence of an evident relationship between lipophilicity and cytotoxicity seem to suggest that all these experimental parameters are probably smoothed out during the 3-day cytotoxicity experiments and do not strongly affect the half-maximal inhibitory concentrations. The results of electrophoretic studies indicate that different kinds of interaction with DNA can be involved in the mode of action of these complexes, with intercalation in double-stranded DNA and stacking on G-quadruplex DNA being strongly implicated in particular for [PtCl2(NH3)(2-aminonaphthalene)].

Copper(II) complexes have been intensely investigated in a variety of diseases and pathological conditions due to their therapeutic potential. The development of these complexes requires a good knowledge of metal coordination chemistry and ligand design to control species distribution in solution and tailor the copper(II) centers in the right environment for the desired biological activity. Herein we present the synthesis and characterization of two ligands HL1 and H2L2 containing a phenanthroline unit (phen) attached to the amino group of histidine (His). Their copper(II) coordination properties were studied using potentiometry, spectroscopy techniques (UV–vis and EPR), mass spectrometry (ESI-MS) and DFT calculations. The data showed the formation of single copper complexes, [CuL1]+ and [CuL2], with high stability within a large pH range (from 3.0 to 9.0 for [CuL1]+ and from 4.5 to 10.0 for [CuL2]). In both complexes the Cu2+ ion is bound to the phen unit, the imidazole ring and the deprotonated amide gr...

Aiming to explore alternative mechanisms of cellular uptake and cytotoxicity, we have studied a new family of copper(II) complexes (CuL1-CuL4) with bis(thiosemicarbazone) (BTSC) ligands containing pendant protonable cyclic amines (morpholine and piperidine). Herein, we report on the synthesis and characterization of these new complexes, as well as on their biological performance (cytotoxic activity, cellular uptake, protein and DNA binding), in comparison with the parental CuIIATSM (ATSM = diacetyl-bis(N4-methylthiosemicarbazonate) complex without pendant cyclic amines. The new compounds have been characterized by a range of analytical techniques including ESI-MS, IR spectroscopy, cyclic voltammetry, reverse-phase HPLC and X-ray spectroscopy. In vitro cytotoxicity studies revealed that the copper complexes are cytotoxic, unlike the corresponding ligands, with a similar potency to that of CuATSM. Unlike CuATSM, the new complexes were able to circumvent cisplatin cross-resistance. The presence of the protonable cyclic amines did not lead to an enhancement of the interaction of the complexes with human serum albumin or calf thymus DNA. However, CuL1–CuL4 showed a remarkably augmented cellular uptake compared with CuATSM, as proved by uptake, internalization and externalization studies that were performed using the radioactive congeners 64CuL1-64CuL4. The enhanced cellular uptake of CuL1-CuL4 indicates that this new family of CuIIBTSC complexes deserves to be further evaluated in the design of metallodrugs for cancer theranostics.

Tridentate pyrazole-containing ligands of the Schiff base type, SalPz — HL 1 , Cl 2 SalPz — HL 2 and I 2 SalPz — HL 3 , were used to prepare a series of new Cu(II) complexes (CuSalPz — 1 , CuCl 2 SalPz — 2 and CuI 2 SalPz — 3 ). These new complexes have been studied by different analytical techniques (electrospray ionization mass spectrometry (ESI-MS), elemental analysis, FT-IR and EPR). The spectroscopic properties of 1 – 3 are consistent with the formation of Cu(II) complexes coordinated by monoanionic and tridentate (N,N,O)-chelators, behaving as monomeric species in aqueous solution, as shown by EPR studies. Crystals of 2 and 3 , obtained by slow concentration of methanolic solutions of the compounds, were also analyzed by X-ray diffraction analysis. The X-ray structural study has shown that 2 crystallized as a dinuclear compound, [Cu 2 (μ-Cl) 2 (Cl 2 SalPz) 2 ], while the solid state structure determined for 3 is best described by monomeric units of [CuCl(I 2 SalPz)] displaying short Cu···Cl intermolecular contacts. The in vitro evaluation of 1 – 3 comprised the study of their DNA-cleaving ability using plasmid DNA and the assessment of their cytotoxic activity against several human cancer cell lines (PC-3 prostate, MCF-7 breast and A2780 and A2780cisR-ovary). The studies with plasmid DNA have shown that 2 and 3 induce extensive DNA cleavage in the presence of different additives. The cytotoxic activity of 2 and 3 is comparable to the one presented by cisplatin, with the exception of the A2780 cell line where cisplatin is more active. It has been found that the introduction of halogen substituents in the phenolate rings of the chelators enhanced the cytotoxicity of the respective Cu(II) complexes.

The molybdenum(VI) binding properties of a series of 3-hydroxy-2-methyl-4-pyridinones (3,4-HP), namely bis(3,4-HP)chelators, were investigated in solution and as solids; in particular, the complexation of MoVI with iminobis{acetyl[1-(3′-aminopropyl)-3-hydroxy-2-methyl-4-pyridinone]}, IDAPr(3,4-HP)2, and ethylenediamino-N,N′-bis(methylcarboxyl)-N,N′-bis{acetyl[1-(3′-aminopropyl)-3-hydroxy-2-methyl-4-pyridinone]}, EDTAPr(3,4-HP)2, as well as three model compounds: 1,2-dimethyl-3-hydroxy-4-pyridinone (DFP), iminodiacetic acid (IDA), and ethylenediaminodiacetic acid (EDDA). Solution studies show that the complexation occurs at low pH values by the coordination modes of the hydroxypyridinone moiety, and with the formation of bischelated MoO22+ species. As the pH is increased, hydrolytic processes start with the formation of monochelated MoO3 complexes and is followed by the total recovery of the free ligand, at pH > 8 for most of the systems, but at pH 10 for EDTAPr(3,4-HP)2. The obtained solid molybdenum complexes showed evidence for the presence of mononuclear species in solution, whereas the dinuclear species, (MoO2)2EDTAPr(3,4-HP)2, were only detected for the EDTA derivative. Aqueous solution studies were performed by pH–potentiometry, spectrophotometry, 1H NMR and 17O NMR spectroscopy, and solid complex species were characterized by elemental analysis, IR spectroscopy, and ESI-MS.(© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007)

The formation of quadruple-stranded DNA induced by planar metal complexes has particular interest in the development of novel anticancer drugs. This is especially relevant for the inhibition of telomerase, which plays an essential role in cancer cell immortalization and is overexpressed in ca. 85-90% of cancer cells. Moreover, G-quadruplexes also exist in other locations in the human genome, namely oncogene promoter regions, and it has been hypothesized that they play a regulatory role in gene transcription. Herein we report a series of new anthracene-containing terpyridine ligands and the corresponding Cu(II) and Pt(II) complexes, with different linkers between the anthracenyl moiety and the terpyridine chelating unit. The interaction of these ligands and metal complexes with different topologies of DNA was studied by several biophysical techniques. The Pt(II) and Cu(II) complexes tested showed affinity for quadruplex-forming sequences with a good selectivity over duplex DNA. Importantly, the free ligands do not have significant affinity for any of the DNA sequences used, which shows that the presence of the metal is essential for high affinity (and selectivity). This effect is more evident in the case of the Pt(II) complexes. Moreover, the presence of a longer linker between the chelating terpyridine unit and the anthracene moiety enhances the interaction with G-quadruplex-forming sequences. We further evaluated the ability of the Cu(II) complexes to interact with, and stabilize G-quadruplex containing regions in oncogene promoters via a polymerase stop assay. These studies indicated that the metal complexes are able to induce G-quadruplex formation and stop polymerase activity.

A new tetradentate ligand having two 3-hydroxy-4-pyridinonate moieties and one carboxylate group appended to a cyclohexane backbone is reported, together with the corresponding bidentate hydroxypyridinonate pendant arm. The synthesis and characterization of these ligands, as well as their complexation properties towards the first transition series of metal ions M(II) (M=Cu, Ni, Zn) in aqueous solution, are described and discussed herein. The tetradentate ligand forms quite stable complexes with this series of metal ions and in solution, it is proved to be a much more effective chelator than the corresponding bidentate derivative. The stability constants of the complexes formed with this set of bivalent ions follow the Irving–Williams order Ni Zn. The carboxylic group has no interaction with the metal centre, thus remaining free for potential interaction with biological ligands (e.g. proteins).

New [PtCl(pz*NN)](n+) complexes anchored by pyrazolyl-diamine (pz*NN) ligands incorporating anthracenyl or acridine orange DNA-binding groups have been synthesized so as to obtain compounds that would display synergistic effects between platination and intercalation of DNA. Study of their interaction with supercoiled DNA indicated that the anthracenyl-containing complex L(2) Pt displays a covalent type of binding, whereas the acridine orange counterpart L(3) Pt shows a combination of intercalative and covalent binding modes with a strong contribution from the former. L(2) Pt showed a very strong cytotoxic effect on ovarian carcinoma cell lines A2780 and A2780cisR, which are, respectively, sensitive to and resistant to cisplatin. In these cell lines, L(2) Pt is nine to 27 times more cytotoxic than cisplatin. In the sensitive cell line, L(3) Pt showed a cytotoxic activity similar to that of cisplatin, but like L(2) Pt was able significantly to overcome cisplatin cross-resistance. Cell-uptake studies showed that L(2) Pt accumulates preferentially in the cytoplasm, whereas L(3) Pt reaches the cell nucleus more easily, as clearly visualized by time-lapse confocal imaging of live A2870 cells. Altogether, these findings seem to indicate that interaction with biological targets other than DNA might be involved in the mechanism of action of L(2) Pt because this compound, despite having a weaker ability to target the cell nucleus than L(3) Pt, as well as an inferior DNA affinity, is nevertheless more cytotoxic. Furthermore, ultrastructural studies of A2870 cells exposed to L(2) Pt and L(3) Pt revealed that these complexes induce different alterations in cell morphology, thus indicating the involvement of different modes of action in cell death. published

A series of four Pt(II) complexes anchored by bidentate or tridentate pyrazolyl-alkylamine chelators bearing different substituents at the azolyl rings has been prepared with the aim to assess their interest in the design of novel anticancer drugs. All complexes have been fully characterized by classical analytical methods and three of them were characterized also by X-ray diffraction analysis. Their solution behavior, together with lipophilicity measurements, cell uptake, antiproliferative properties, DNA interaction have been evaluated. Albeit all the complexes were less active than cisplatin on ovarian carcinoma A2780 cell line, greatly retained their activity in the cisplatin-resistant A2780cisR cell line and presented a lower resistance factor compared to cisplatin. Moreover, the Pt(II) complexes under investigation were less prone to undergo deactivation by glutathione, believed to be the major cellular target of cisplatin that inactivates the drug by binding to it irreversibly.

The study of chelating compounds is very important to solve problems related to human metal overload. 3-Hydroxy-3-pyridinones (HP), namely deferiprone, have been clinically used for chelating therapy of Fe and Al over the last decade. A multi-disciplinary search for alternative molecules led us to develop poly-(3-hydroxy-4-pyridinones) to increase metal chelation efficacy. We present herein a complexation study of a new bis-(3-hydroxy-4-pyridinone)-EDTA derivative with a set of M(3+) hard metal ions (M = Fe, Al, Ga), as well as Zn(2+), a biologically relevant metal ion. Thus a systematic aqueous solution equilibrium study was performed using potentiometric and spectroscopic techniques (UV-Vis, NMR methods). These set of results enables the establishment of specific models as well as the determination of thermodynamic stability constants and coordination modes of the metal complexes. The results indicate that this ligand has a higher affinity for chelating to these hard metal ions than deferiprone, and that the coordination occurs mostly through the HP moieties. Furthermore, it was also found that this ligand has a higher selectivity for chelating to M(3+) hard metal ions (M = Fe, Al, Ga) than Zn(2+).

The DNA cleavage activity of several beta-diketonate vanadyl complexes is examined. Vanadyl acetylacetonate, V(IV)O(acac)(2), 1, shows a remarkable activity in degrading plasmid DNA in the absence of any activating agents, air and photoirradiation. The cleaving activity of several related complexes V(IV)O(hd)(2) (2, Hhd=3,5-heptanedione), V(IV)O(acac-NH(2))(2) (3, Hacac-NH(2)=acetoacetamide) and V(IV)O(acac-NMe(2))(2) (4, Hacac-NMe(2)=N,N-dimethylacetoacetamide) is also evaluated. It is shown that 2 exhibits an activity similar to 1, while 3 and 4 are much less efficient cleaving agents. The different activity of the complexes is related to their stability towards hydrolysis in aqueous solution, which follows the order 1 approximately 23 approximately 4. The nature of the pH buffer was also found to be determinant in the nuclease activity of 1 and 2. In a phosphate buffered medium DNA cleavage by these agents is much more efficient than in tris, hepes, mes or mops buffers. The reaction seems to take place through a mixed mechanism, involving the formation of reactive oxygen species (ROS), namely OH radicals, and possibly also direct cleavage at phosphodiester linkages induced by the vanadium complexes.

3-Hydroxy-4-pyridinones (3,4-HP) are well known iron-chelators with applications in medicinal chemistry, mainly associated with their high affinity towards trivalent hard metal ions (e.g. M(3+), M=Fe, Al, Ga) and use as decorporating agents in situations of metal accumulation. The polydenticity and the extra-functionality of 3,4-HP derivatives have been explored, aimed at improving the chelating efficacy and the selectivity of the interaction with specific biological receptors. However, the ideal conjugation of both features in one molecular unity usually leads to high molecular weight compounds which can have crossing-membrane limitations. Herein, a different approach is used combining a arylpiperazine-containing bis-hydroxypyridone (H(2)L(1)) with a biomimetic mono-hydroxypyridinone, ornithine-derivative (HL(2)), to assess the potential coadjuvating effect that could result from the administration of both compounds for the decorporation of hard metal ions. This work reports the results of solution and in vivo studies on their chelating efficacy either as a simple binary or a ternary system (H(2)L(1):HL(2):M(3+)), using potentiometric and spectrophotometric methods. The solution complexation studies with Fe(III) indicate that the solubility of the complexes is considerably increased in the ternary system, an important feature for the metal complex excretion, upon the metal sequestration. The results of the in vivo studies with (67)Ga-injected mice show differences on the biodistribution profiles of the radiotracer, upon the administration of each chelating agent, that are mainly ascribed to the differences of their extra-functional groups and lipo/hydrophilic character. However, administration of both chelating agents leads to a more steady metal mobilization, which may be attributed to an improved access to different cellular compartments.

Iron (Fe) overload diseases, such as beta-thalassemia (thal) major and hemochromatosis, have been treated for several decades by chelating therapy with desferrioxamine (DFO). However, drawbacks associated with that drug led to the development of new chelating drugs. The 3-hydroxy-4-pyridinones emerged as highly effective Fe chelators, and deferiprone (L1) has been approved as a Fe chelating drug. The most recent strategy for Fe overload problems is based on the replacement of monotherapies by a combination therapy with both chelators. Following a similar chelating strategy, we present herein the results of animal tests with a combination of two different hydroxypyridinone-based chelators. Both are of the 3-hydroxy-4-pyridinone (HP) type, but with one and two HP chelating units, and extra functional groups to account for differentiation in their physicochemical and biological properties, namely chelating efficacy and bioavailability. Animal studies have shown that the simultaneous administration of this pair of HP chelators, under appropriate proportion, to metal-loaded mice, could speed up metal excretion. This may be rationalized by adjuvant and eventual synergistic effects, due to complementary accessibility of each chelator to different cellular compartments.

The 3-hydroxy-4-pyridinones are chelating agents of current interest due to their high affinity for hard metal ions and potential clinical applications as metal-decorporation agents. A new bis-(3-hydroxy-4-pyridinone) derivative of EDTA have been developed, and herein we describe the results of solution studies to determine the protonation constants and the partition coefficient. Biodistribution studies, performed with 67Ga-overload mice, showed rapid clearance of the radiotracer from the body, thus indicating that the new ligand should be a quite effective agent for the in vivo aluminium removal.

A new bis(3-hydroxy-4-pyridinone) derivative of iminodiacetic acid, imino-bis(acetyl(1-(3′-aminopropyl)-3-hydroxy-2-methyl-4-pyridinone)), IDAPr(3,4-HP)2, has been prepared and studied in its interaction with a set of hard metal ions. This tetradentate ligand presents a much higher chelating efficiency for trivalent hard metal ions (Fe, Ga, Al) than the monodentate derivative Deferriprone, namely at the diluted conditions prevailing in physiological conditions and at low clinical doses. A similar behaviour was also observed for the complexation with Zn(II) but at a significantly lower extent. This compound presents a moderate hydrophilic character at physiological pH (logD = −1.72). In vivo assays showed much more rapid clearance of 67Ga from most tissues of metal-loaded mice than the drug Deferriprone and the radioactivity excretion occurs mostly through the kidneys. Therefore, results from in vitro and in vivo studies indicated good perspectives for this compound to be a potential decorporating agent for hard metal ions in overload situations without depletion of essential metal ions such as zinc.

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© The Royal Society of Chemistry 2010, New pyrazolyl-diamine ligands with acridine derivatives at the 4-position of the pyrazolyl ring were synthesized and characterized (L1 and L2). Coordination towards the fac-[M(CO)3]+ (M = Re, 99mTc) led to complexes fac-[M(CO)3(κ3-L)] (L = L1: M = Re1, Tc1; L = L2: M = Re2, Tc2). The interaction of the novel pyrazolyl-diamine ligands (L1 and L2) and rhenium(I) complexes (Re1 and Re2) with calf thymus DNA (CT-DNA) was investigated by a variety of techniques, namely UV-visible , fluorescence spectroscopy and circular and linear dichroism . Compounds L1 and Re1 have moderate affinity to CT-DNA and bind to DNA by intercalation, while L2 and Re2 have a poor affinity for CT-DNA. Moreover, LD measurements showed that L1 and Re1 act as perfect intercalators . By confocal fluorescence microscopy we found that L1 and Re1 internalize and localize in the nucleus of B16F1 murine melanoma cells . The congener Tc1 complex also targets the cell nucleus exhibiting a time-dependent cellular uptake and a fast and high nuclear internalization (67.2% of activity after 30 min). Plasmid DNA studies have shown that Tc1 converts supercoiled (sc) puc19 DNA to the open circular (oc) form., Teresa Esteves and Sofia Gama thank the FCT for a doctoral and postdoctoral research grants (SFRH/BD/29154/2006 and SFRH/BPD/29564/2006, respectively). COST Action D39 is also acknowledge. The QITMS instrument was acquired with the support of the Programa Nacional de Reequipamento Científico (Contract>REDE/1503/REM/2005-ITN) of Fundação para a Ciência e a Tecnologia and is part of RNEM - Rede Nacional de Espectrometria de Massa.

Dissertação de Mestrado em Política Social O consumo de drogas e as suas consequências constituem um drama humano e social onde se encontram envolvidos os consumidores, o seu seio familiar, laboral e social. Sabemos que a fase da reinserção social é parte integrante do processo de recuperação da toxicodependência, como tal, o presente trabalho pretende contribuir para a compreensão da importância desta etapa e relacionar a mesma com a questão do género. Este estudo ambiciona, também, salientar a necessidade de se ter em consideração, em processo de reinserção, que qualquer que seja a pessoa a que se destina, esta é um ser pleno de direitos que está envolvido num meio, pertencente a vários subsistemas. Portanto, cada processo de reinserção é único pois cada indivíduo tem necessidades diferenciadas. Em termos científicos, o objetivo será contribuir para uma melhor compreensão do flagelo que é a toxicodependência e todo o seu processo de recuperação. We can see the use of drugs and the consequences of this addiction, as a human and social drama, that includes the drugs addicts, his family and his social and work relationship. As we know, the social reinsertion stage is one part of the recuperation process of the drug addiction, so, the main objective of this work is try to elucidate to the importance of this stage, and make a connection with the genre question. This study try as well, to demonstrate the necessity to considerate , along of the reinsertion process, that any person involved have all social rights and is involved in a determined social reality and is a part of various subsystems. So, any reinsertion process is unique, because the addicts have many different necessities. In scientific terms, the objective is to give an contribute to a better perception of the scourge of the drug addiction and is recuperation process.