Submitted by Luciana Ferreira (lucgeral@gmail.com) on 2014-12-01T11:55:38Z No. of bitstreams: 2 Dissertação - Bruna dos Santos Rodrigues - 2013.pdf: 1797836 bytes, checksum: b56e1b3bdd63eeb9040d2ef050f157f8 (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2014-12-04T14:18:47Z (GMT) No. of bitstreams: 2 Dissertação - Bruna dos Santos Rodrigues - 2013.pdf: 1797836 bytes, checksum: b56e1b3bdd63eeb9040d2ef050f157f8 (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Made available in DSpace on 2014-12-04T14:18:47Z (GMT). No. of bitstreams: 2 Dissertação - Bruna dos Santos Rodrigues - 2013.pdf: 1797836 bytes, checksum: b56e1b3bdd63eeb9040d2ef050f157f8 (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) Previous issue date: 2013-08-30 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / The death mechanisms induced by a new synthetic compound (4-FTC) in adenocarcinoma prostate cells (PC-3) and its toxicity were investigated in this study. PC-3 cells cytotoxity was evaluated by MTT reduction assay. The mechanisms involved in PC-3 death and cell cycle were investigated by flow cytometry and colorimetric assays. The compound toxicity was analized by cytotoxicity of mononuclear cells (MTT reduction assay) and 3T3 cells (neutral red uptake assay), myelotoxicity, haemolytic activity and acute oral toxicity. 4-FTC has concentration dependent cytotoxic activity in PC-3 cells, and 184,6 μM IC50. Investigation of death mechanisms indicated death by apoptosis, because of the significant increase in phosphatidylserine externalization (109,83%), loss of mytochondrial membrane potential (41,96%), significant increase of DNA fragmentation (284,02%) and capases 3/7 and 9 activity increase, 13,12% and 12,8%, respectively. Furthermore, the treatment of PC-3 cells wih 4-FTC did not induce the reactive oxygen species production, as well as, the induction of acid autophagic vesicles generation and did not change the cell cycle significantly. Althought 4-FTC was able to modulate the expression of some proteins that regulate cell cycle, incresead the expression of p53, p21 and p27. Thus, the results suggests that 4-FTC induced PC-3 death by apoptosis dependent by mitochondrial pathway activation. In toxicity evaluation, 4-FTC presented 52,86 μM and 19,63 μM IC50 to mononuclear and 3T3 cells, respectively; 27,35 μM IC50 to hematopoietic precursors; low acute oral toxicity, classified in GHS category 5, and not significant haemolytic activity. / Neste trabalho, investigaram-se os mecanismos de morte induzidos por um novo composto sintético, 4-fluorbenzaldeidotiossemicarbazona (4-FTC), nas células de adenocarcinoma prostático PC-3 e alguns parâmetros da toxicidade desse composto. A citotoxicidade nas células PC-3 foi avaliada pelo método de redução do MTT, método colorimétrico para avaliar a viabilidade celular. A investigação dos mecanismos de morte induzidos foi realizada por citometria de fluxo e ensaio colorimétrico. A toxicidade do composto foi avaliada pela citotoxicidade em células mononucleares pelo método de redução do MTT, citotoxicidade em células 3T3 pelo método de incorporação do vermelho neutro, mielotoxicidade, potencial hemolítico e toxicidade oral aguda. Os resultados obtidos mostram atividade citotóxica concentração dependente, com CI50 de 184,6 μM para PC-3. A investigação dos mecanismos de morte induzidos indicou morte por apoptose, pois houve aumento significativo da externalização da fosfatidilsserina (109,83%), perda do potencial de membrana mitocondrial em 41,96%, aumento significativo da fragmentação de DNA (284,02%) e aumento de caspases 3/7 e 9, em 13,12% e 12,8%, respectivamente. Além disso, não induziu a produção de espécies reativas de oxigênio, bem como, a formação de vesículas autofágicas ácidas e não alterou o perfil do ciclo celular de forma significativa. Embora tenha modulado a expressão de proteínas reguladoras do ciclo celular, aumentando a expressão de p53, p21 e p27. Assim, pode-se sugeririr que o 4-FTC induz morte por apoptose por meio de mecanismos de ativação dependentes da via mitocondrial em células PC-3. Na avaliação da toxicidade, o 4-FTC apresentou concentração inibitória 50% (CI50) de 52,86 μM e 19,63 μM para células mononuclerares e células 3T3, respectivamente; CI50 de 27,35 μM para precursores hematopoiéticos; baixa toxicidade oral aguda, sendo classificado na categoria 5 e baixo potencial hemolítico.

A series of dioxidomolybdenum(VI) complexes, [MoVIO2L1–6] (1–6) and [MoVIO2L1–6(solv)] (1a–6a) {where solv (solvent) = DMSO (1a, 3a, 5a and 6a) and H2O (2a and 4a)} have been synthesized using thiosemicarbazone ligands, H2L1–6. Furthermore, six monooxidomolybdenum(IV) complexes [MoIVOL1–6(N-N)] (7–12) {where co-ligand (N-N) = 2,2′-bipyridine (bipy) (7, 10 and 11) and 1,10-phenanthroline (phen) (8, 9 and 12)} have also been synthesized from the corresponding Mo(VI) precursors, [MoVIO2L1–6] (1–6) by oxygen atom transfer (OAT) reaction. Complexes have been characterized by conventional methods, including X-ray crystallography, and DFT (density functional theory) calculations. OAT reactivity of Mo(VI) and Mo(IV) complexes have been successfully established through the formation of OPPh3 and Me2S. These OAT products have been characterized by 31P NMR (OPPh3), UV–Vis spectroscopy and GC–MS (Me2S) and DFT simulations supported this finding through the prediction of ΔGtotsol for the reaction of oxygen atom transfer. DFT methods suggested that the oxygen atom transfer from [MoVIO2L] species to PPh3 to give [MoIVOL(bipy)] and from DMSO to [MoIVOL(bipy)] to yield [MoVIO2L] is strongly favored, whereas the formation of μ-oxido dimer [MoV2O3L2], is much less probable.

No description available.

<p>Opisane su sinteze, fizičko-hemijske i strukturne karakteristike kompleksa Cu(II), Co(II), Co(III), Ni(II), Fe(III), Cr(III) Zn(II) i Cd(II) sa PLSC, PLTSC i PLITSC. Dobijene su dve nove protonovane forme liganda PLSC, kao i od ranije poznati neutralna forma PLSC i protonovana forma PLTSC ali u vidu monokristala pogodnih za rentgeno-strukturnu analizu i 37 kompleksa, od čega je 12 okarakterisano rentgeno-strukturnom analizom. U zavisnosti od pH sva tri liganda se mogu koordinovati kao neutralni, mono-, odnosno, dianjoni sa uobičajenim ONO, ONS i ONN setom donorskih atoma, respektivno, pri čemu neutralna i monoanjonska forma liganada postoji u obliku zwitter-jona. U slučaju PLTSC a na primerima dva dimerna kompleksa bakra nađen je i ređi tetradentatni ONSO način koordinacije, tj. uče&scaron;će i atoma kiseonika hidroksimetil grupe kao mostovnog atoma. Sva tri liganda sa Cu(II), nezavisno od stehiometrije Cu:ligand, daju mono(ligand) komplekse kvadratno-planarne, kvadratno-piramidalne ili oktaedarske strukture, a sa kobaltom mono- i bis(ligand) oktaedarske komplekse. Dobijenim mono(ligand) kompleksima Zn(II) sa sva tri liganda je pripisana najmanje tetra-, odnosno pentakoordinacija, dok je me&scaron;ovitim PLSC(PLTSC)/NCS i bis(ligand) kompleksima Cr(III), i mono(ligand) kompleksu Fe(III) sa PLTSC pripisuje oktaedarska struktura. Ni(II) sa PLTSC i PLITSC daje mono- i dinuklearne komplekse, dok Cd(II) sa istim ligandima gradi mono(ligand) komplekse. Sva jedinjenja su okarakterisana parcijalnom mikroanalizom, IR-spektrima i konduktometrijskim merenjima, a odabrani kompleksi rentgeno-strukturnom, termogravimetrijskom i toksikolo&scaron;kom analizom.</p> / <p>The work describes the syntheses, physico-chemical, structural and antibacterial characteristics of Cu(II), Co(II), Co(III), Zn(II), Ni(II), Fe(III), Cr(III) and Cd(II) complexes with semi-, thiosemi- and S-methylisothiosemicarbazones. The result of the syntheses was 2 new ligand forms and 37 complexes, of which 12 were characterized by X-ray structural analysis. Depending of the pH, all three ligands can be coordinated either as neutral, monoanion, or dianion, using ONO, ONS and ONN set of donor atoms, respectively, whereby the neutral and monoanionic forms of the ligands exist in the form of zwitter-ions. All three ligands form with Cu(II), irrespective of the stoichiometry Cu:ligand, give mono- and dinuclear complexes of square-planar, square-pyramidal and/or octahedral configuration and with cobalt mono- and bis(ligand) octahedral complexes. The obtained mono(ligand) complexes of Zn (II) with all three of ligands is attributed to the at least tetra-, or pentacoordination, while the mono- and bis(ligand) complexes of Cr(III) with PLSC and PLTSC, and mono (ligand)complex of Fe(III) with PLTSC attributable to octahedral structure. Ni(II) with PLTSC and PLITSC forms mono- and binuclear complexes, and Cd(II) with the same ligands forms mono(ligand) complexes. All the compounds were characterized by partial microanalysis, IR spectra and conductometric measurements, while some selected complexes were examined by X-ray structural, thermogravimetric and toxicological analysis.</p>

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Abnormalities of lipid metabolism often lead to pathologic lipid accumulation in the vessel wall, oxidative and chronic inflammatory sequelae and the formation of atherosclerotic lesions, ultimately leading to clinical events. Low density lipoprotein (LDL) oxidative modification in the vascular wall seems to be a key factor in atherosclerosis development. Following the oxidation hypothesis of atherosclerosis the role of antioxidants has been investigated in a large number of epidemiological, clinical and experimental studies. Therefore, we investigated the antioxidant activity of three compounds of different classes in preventing the low density lipoprotein oxidation. In the guaraná study, we showed that subjects who habitually ingested guaraná (GI) demonstrated lower conjugated dienes production than did subjects who never ingested guaraná (NG; reduction of 27%, p < 0,0014), independent of other variables. However, in the GI group the conjugated dienes production was positively associated with glucose levels. The GI group also showed a total cholesterol level significantly lower than NG group. Also, guaraná demonstrated a high antioxidant activity in vitro, mainly at concentrations of 1 and 5 μg/mL, demonstrated by suppression of CDs and TBARS productions, tryptophan destruction and high TRAP activity. Guaraná, similar to other foods rich in caffeine and catechins such as green tea, has some effect on LDL oxidation that could partially explain the protective effects of this food in cardiometabolic diseases. In the thiosemicarbazone study, salicylaldehyde-4- phenylthiosemicarbazone (SPTS) may have antioxidant activity against Cu2+- and 2,2′- Azobis(2-methylpropionamidine) dihydrochloride (AAPH)-induced LDL and serum oxidation. Additionally, SPTS was effective at preventing tryptophan destruction. SPTS also showed significant total radical-trapping antioxidant activity and could prevent thiobarbituric acid reactive substances (TBARS) formation induced by sodium nitroprusside in different rat tissues and by Cu2+ in human LDL and serum. These results indicate that the antioxidant effect of SPTS is caused by a combination of transition metal chelation and free-radicalscavenging activity. In the organotellurium study, the 2-phenyl-2-tellurophenyl vinylphosphonate (DPTVP) may have antioxidant activity against Cu2+- and 2,2′-azobis(2- methylpropionamidine) dihydrochloride (AAPH)-induced LDL and serum oxidation. Additionally, DPTVP was effective at preventing tryptophan oxidation. DPTVP also showed significant total radical-trapping antioxidant activity and could prevent thiobarbituric acid reactive substance (TBARS) formation induced by Cu2+ in human LDL and serum. Additionally, DPTVP exhibited no toxicity in rat aorta slices. The results presented here indicate that the antioxidant effect of DPTVP is caused by a combination of free-radicalscavenging activity and possible blockade of the copper binding sites of LDL. Considering these preliminary results, we can conclude that the three compounds presented a potential antioxidant activity and could prevent the oxidative modifications of LDL. These data encourage us to evaluate these compounds in in vivo studies and investigate additional properties in preventing the atherogenic process. / Anormalidades do metabolismo lipídico muitas vezes levam ao acúmulo patológico de lipídios na parede arterial, sequelas oxidativas e inflamatórias crônicas e à formação de lesões ateroscleróticas, levando a eventos clínicos. A modificação oxidativa de lipoproteínas de baixa densidade (LDL) na parede arterial parece ser um fator importante no desenvolvimento da aterosclerose. Seguindo a hipótese oxidativa da aterosclerose, o papel dos antioxidantes tem sido investigado em grande número de estudos epidemiológicos, clínicos e experimentais. Sendo assim, neste estudo nós investigamos a atividade antioxidante de três compostos de diferentes classes na prevenção da oxidação da LDL. No estudo com guaraná, nós mostramos que indivíduos que consumiam guaraná habitualmente (GI) apresentaram menor produção de dienos conjugados que os indivíduos que nunca consumiam guaraná (NG; redução de 27%, p < 0,0014), independentemente de outras variáveis. No entanto, no grupo GI a produção de dienos conjugados foi positivamente associada com os níveis de glicose. O grupo GI também apresentou nível de colesterol total significativamente menor comparado ao grupo NG. Além disso, o guaraná apresentou uma grande atividade antioxidante in vitro, principalmente nas concentrações de 1 e 5 μg/mL, demonstrado pela supressão da produção de dienos conjugados e substâncias reativas ao ácido tiobarbitúrico, prevenção da destruição do triptofano e alta atividade scavenger de radicais (TRAP). O guaraná, similar a outros alimentos ricos em cafeína e catequinas como o chá verde, tem alguns efeitos na oxidação da LDL que podem explicar parcialmente os efeitos protetores deste alimento nas doenças cardiometabólicas. No estudo com a tiosemicarbazona, a salicilaldeído-4-feniltiosemicarbazona (SPTS) apresentou atividade antioxidante contra a oxidação de LDL e soro induzidas por Cu2+ e 2,2 -azobis(2- metilpropionamidina) dihidrocloreto (AAPH). Além disso, a SPTS foi efetiva em prevenir a destruição do triptofano. O composto também apresentou significativa atividade scavenger de radicais e pode prevenir a formação de substâncias reativas ao ácido tiobarbitúrico induzidas por nitroprussiato de sódio em diferentes tecidos de ratos e por Cu2+ em LDL e soro humano. Estes resultados indicam que o efeito antioxidante da SPTS é causado pela combinação da atividade quelante do composto e a atividade scavenger de radicais livres. No estudo com o composto orgânico de telúrio, o 2-fenil-2-telurofenil vinilfosfonato (DPTVP) apresentou atividade antioxidante contra a oxidação de LDL e soro humano induzida por AAPH e Cu2+. Além disso, o composto preveniu a oxidação do triptofano e a formação de substâncias reativas ao ácido tiobarbitúrico e mostrou um significativo efeito scavenger de radicais. O DPTVP (20 μM) não apresentou toxicidade quando exposto à fatias de aorta de ratos. Estes resultados indicam que o efeito do DPTVP é resultado de uma combinação da atividade scavenger de radicais do composto e da possibilidade dele bloquear os sítios de ligação de cobre da LDL. Considerando todos os resultados apresentados aqui, podemos concluir que os três compostos apresentam um grande potencial antioxidante e podem prevenir as modificações oxidativas da LDL. Esses dados nos encorajam para avaliar esses compostos em estudos in vivo e investigar novas propriedades que possam prevenir o processo aterogênico.

Use of metal based anticancer medication began with the clinical approval of cisplatin in 1978. Research led to the development of six platinum based drug candidates which are in use around the world. However there is a great need to develop better treatment strategies. The present work entitled “Cytotoxicity of Metal Based Anticancer Active Complexes and Their Targeted Delivery Using Nanoparticles” is an effort to prepare cytotoxic metal complexes based on platinum(IV) and copper(II) and deliver them selectively to cancer cells using a targeting ligand, biotin, with two different delivery vehicles, viz. PEGylated polyamidoamine dendrimer (PAMAM) and gold nanoparticles (AuNPs). Chapter 1 provides a brief introduction to cancer and its characteristic features, followed by a short description about different treatment modalities in clinical practice. An account of the development of anticancer drugs starting from purely organic drugs to the field of metal based anticancer drugs is discussed. An overview of the available targeting strategies are discussed with specific examples. The section ends with the scope of the present work. Platinum based anticancer drugs currently in use contain platinum in the +2 oxidation state. These drugs showed side effects and are often ineffective against resistant cells, especially in the latter stages of treatment. A recent focus of metal based anticancer drug research is the development of platinum(IV) systems which shows promise to have greater activity in cancer cells in a reducing environment. Reported platinum(IV) dual drugs contain the components of “cisplatin” or an analogue along with an active organic drug. But there are no known dual drugs based on platinum(IV) that would generate a cytotoxic metal complex along with cisplatin. In Chapter 2, a bimetallic dual drug (M4) (Figure 1), the first of its kind, with components of cisplatin and copper bis(thiosemicarbazone) has been prepared (Figure 1). The components and the bimetallic complex were characterized using several spectroscopic techniques. The dual drug M4 was found to be highly cytotoxic (IC50 1.3 M) against HeLa cells and was better than cisplatin (IC50 6.8 M). The bimetallic complex turned out to be better than the mixture (IC50 7.2 M) of individual drugs which indicated possible synergism of the released cisplatin and the copper bis(thiosemicarbazone) from the dual drug. Figure 1: Structure of the platinum(IV) and copper bis(thiosemicarbazone) complexes. A novel approach towards conjugation of platinum(IV) drugs to a carrier has been developed using a malonate moiety (Figure 2). The bis(butyric acid) complex, Pt(NH3)2(OCOC3H7)2Cl2 (M1), was taken as model complex to demonstrate the conjugation strategy. The complex M4 was also conjugated to the partially PEGylated 5th generation PAMAM dendrimers. Figure 2: Schematic representation of the platinum(IV) drug conjugated PAMAM dendrimer. The cytotoxicity of M4 was reduced to a small extent on conjugation to the dendrimer. In the presence of 5 mM sodium ascorbate as a reducing agent, sustained release (40 %) of the drug was shown to occur over a period of 48 h by the drug release study. The reduction in cytotoxicity of the dendrimer conjugates could be due to incomplete release of the active drug. Unfortunately, no enhanced activity was observed with the additional targeting ligand, biotin. The drug uptake study revealed that the dendrimer conjugates were successful in entering cancer cells. There was no preferential uptake with biotin conjugated dendrimers which explained the similar cytotoxicity of dendrimer conjugates with and without biotin. Different delivery vehicles showed varied efficiency in delivering the pay load (drugs) to the cancer site. In this connection, PEGylated gold nanoparticles have shown good promise as a drug delivery vehicle. In Chapter 3, M1 and M4 are both conjugated to malonate functionalized PEGylated gold nanoparticles (30 nm). Biotin was also attached to the AuNPs for targeting HeLa cells. Figure 3: Schematic representation of the platinum(IV) drug and biotin conjugated AuNPs. The AuNPs were highly stable in water without agglomeration. There was no shift in the Surface Plasmon Resonance (SPR) band after conjugation of the drug molecules and targeting ligands. TEM images and DLS measurements showed there was no change in particle size. Drug conjugated AuNPs were also very stable in high salt concentrations as well as over a large range of pH. AuNPs with M1 were found to be less cytotoxic than the parent drug. Biotinylated AuNPs with M1 were more potent than non-biotinylated nanoparticles and increased cytotoxicity (35 %) was observed with biotin conjugation. Surprisingly, the enhanced activity of biotinylated AuNPs could not be correlated to the drug uptake study. The cytotoxicity of the bimetallic dual drug containing AuNPs were about 10-fold less and no increased activity was observed with the biotinylated conjugates. The reduced activity of AuNPs with the bimetallic drug was due to incomplete release from the AuNPs (20 % release after 48 h). But the release kinetics was very slow and sustained which might increase in vivo activity. The unexpected lower activity of biotinylated conjugates with copper bis(thiosemicarbazone) was suggestive of interference between bis(thiosemicarbazone) complex and the biotin receptor resulting in reduced drug uptake. Copper bis(thiosemicarbazone) complexes hold very good promise as a class of non-platinum anticancer drug candidates. However, they lack selectivity towards malignant cells. Recently, CuATSM has shown hypoxia selectivity and very good cytotoxicity resulting in 64CuATSM being used in advanced stages of clinical trials for imaging hypoxic cells. In Chapter 4, a copper bis(thiosemicarbazone) complex analogous to Cu(ATSM) with a redox active cleavable disulfide linker and a terminal carboxylic acid group (CuATSM-SS-COOH) was synthesised and characterised spectroscopically. The complex was highly cytotoxic and has an IC50 value (6.9 M) similar to that of cisplatin against HeLa cells. The complex was conjugated to PEGylated gold nanoparticles by amide coupling between the acid group from the drug molecule and the amine on the AuNPs (20 nm) for smart drug delivery. The gold nanoparticles were decorated with biotin for targeted delivery to the HeLa cells. Figure 4: Schematic representation of the CuATSM-SS-COOH and biotin decorated AuNPs. The CuATSM-SS-COOH was insoluble in water but conjugation to PEGylated gold nanoparticles made it water soluble. The drug molecules and biotin conjugated AuNPs were highly stable which was confirmed by TEM and DLS measurements. Similar to the study described in the previous chapter, these AuNPs were also stable in a wide range of pH and salt concentrations. In vitro glutathione (GSH) triggered release study demonstrated substantial release of the cytotoxic agent from the AuNPs (60 %) over a period of 48 h. In vitro cell viability study with HeLa cells showed reduced cytotoxicity (IC50 15 M) of AuNPs with and without biotin containing drug conjugates relative to the parent copper complex (IC50 6.9 M). The reduction of the cytotoxicity correlated well with the released amount of the active drug from the nanoconjugates over the same time period. In vivo studies demonstrated the effectiveness of these nanoparticle carriers as suitable vehicles as they exhibited nearly four-fold reduction of tumor volume without significant loss in body weight. Moreover, the biotin targeted nanoparticle showed significant (p < 0.5) reduction in tumor volume compared to the non-targeted gold nanoparticles. Thus, this smart linking strategy Can be extended to other cytotoxic complexes that suffer from non-specificity, low aqueous solubility and toxicity. Multinuclear anticancer active complexes do not act in the same way as that of their corresponding mononuclear analogues. In the case of multinuclear platinum complexes, the activity not only depends on the active moiety but also on the spacer length between the moieties. In Chapter 5, a series of multinuclear copper bis(thiosemicarbazone) complexes were prepared and characterised using different techniques. Figure 5: General structures of binuclear copper bis(thiosemicarbazone) complexes. All the complexes showed redox activity and have a very high negative reduction potential, i.e. these compounds would not be easily reduced in the biological medium and would remain as copper(II) species. As the concentration of the reducing agents are more within cancer cells, once these complexes are inside cells they would be reduced to Cu(I). These compounds were shown to be highly lipophilic from the large log P values. Unfortunately, these binuclear complexes were less active than similar mononuclear complexes. One possible reason for the reduced cytotoxicity of these complexes could be adherence of the complexes to the cell membrane due to the high lipophilicity of these complexes. Out of five different methylene spacers between two bis(thiosemicrarbazone) moieties, the complex with a three carbon spacer was shown to be the most active against HeLa cells. The complexes with five and six methylene spacers turn out to be noncytotoxic. Further experiments are necessary to reveal the mechanism of action in these complexes. In summary, bimetallic complexes can be very active and may be a way of overcoming drug resistance in platinum based therapy. A dual drug can be delivered using a malonate moiety and a disulfide linker. Gold nanoparticles are good delivery vehicles for these dual drugs and show great potential for improvement and translation to the next stage. (For figures pl refer the abstract pdf file)