A tuberculose (TB) é uma doença infectocontagiosa causada principalmente pelo Mycobacterium tuberculosis (Mtb). A TB era uma doença considerada em vias de extinção, porém sua incidência voltou a aumentar devido a diversos fatores, como o fluxo migratório dos chamados \"Países Reservatórios\" e a pandemia da AIDS. Em 2013, a OMS estimou 9 milhões de novos casos de TB e 1,5 milhões de mortes. A multirresistência aos quimioterápicos disponíveis justifica a procura por novos tuberculostáticos. Face ao exposto, o trabalho objetivou a síntese de potenciais inibidores da maltosiltransferase (GlgE), um novo e promissor alvo contra o Mtb, utilizando Planejamento com base na Estrutura do Alvo Molecular -- Structure-Based Drug Design (SBDD). A comparação das simulações de dinâmica molecular do sistema apo da GlgE e em complexo com a maltose-1-fosfato (M1P) mostrou as mudanças estruturais, em nível atômico, decorrentes da ligação com seu substrato. Levando em consideração estes dados, as simulações de docking dos potenciais inibidores revelaram que análogos glicosídicos e maltosídicos triazólicos ligados a grupos aromáticos substituídos possuem melhores perfis de interação com o sítio ativo da GlgE e, por essa razão, foram escolhidos para síntese. Assim, sintetizaram-se análogos glicosídicos e maltosídicos através da click chemistry, obtendo-se rendimentos variados. Posteriormente, realizaram-se ensaios enzimáticos e microbiológicos no Institute for Tuberculosis Research (ITR), University of Illinois at Chicago, USA. Apesar de os ensaios enzimáticos estarem em andamento, os compostos ensaiados apresentaram atividades que variaram de moderada a fraca, frente à cepa H37Rv. Dois compostos glicosídicos exibiram valores de Concentração Inibitória Mínima (CIM) inferiores a 60 µM, sem efeito citotóxico em células VERO em concentrações superiores a 250 µM. Tais resultados indicam que o análogo glicosídico triazólico aromático, como o NDS 112, pode ser utilizado como protótipo interessante para o planejamento de novas séries de compostos capazes de atuar como tuberculostáticos / Tuberculosis (TB) is an contagious infectious disease mainly caused by Mycobacterium tuberculosis (Mtb). TB was a disease considered in extinction, but its incidence has risen again due to several factors, such as the migration from \"Reservatoir Countries\" and AIDS pandemic. In 2013, WHO estimated 9 million new TB cases and 1.5 million deaths. Multidrug resistance to drugs available in chemotherapy validates the search for new TB drugs. In this context, this work aimed to the design and synthesis of potential inhibitors of maltosyltransferase (GlgE), a new and promissor target against Mtb using Structure-Based Drug Design. Comparison between molecular dynamic simulations of GlgE apo system and in complex with maltose-1-phosphate (M1P) showed important structural changes at molecular level due to substrate binding. Considering these data, docking simulations of potential inhibitors revealed that glucoside and maltoside triazoles analogues bound to substituted aromatic groups have better interaction profiles with the GlgE active site and for this reason they were chosen for synthesis. Thus, glucoside and maltoside triazole analogues were synthetized through click chemistry with different yields. Then, enzymatic and microbiological assays were performed at Institute for Tuberculosis Research (ITR), University of Illinois at Chicago, USA. Although the enzymatic assays are ongoing, componds were tested against H37Rv strains and they have shown activity ranging from moderate to weak. Two compounds presented Minimal Inhibitory Concentration (MIC) values below 60 µM, without cytotoxic effect in VERO cells in concentrations higher than 250 µM. These results indicate that glucosidic aromatic triazoles analogues, such as NDS 112, can be used as a prototype for design new series of compounds capable of acting as antituberculosis agents

Le développement de nouveaux antibiotiques est un enjeu majeur de santé publique. Etant donné, le fort potentiel des aminoglycosides en tant qu’antibiotique, ces composés ont attisé l’intérêt de plusieurs groupes de recherche. Cependant, leur usage est encore très limité, malgré leur ancienneté, du fait de leur toxicité et du développement toujours croissant des mécanismes de résistances aux aminoglycosides. Afin de mieux appréhender les problèmes inhérents à leur utilisation, il est crucial de mieux comprendre leur action sur les différentes cibles cellulaires, et d’étudier leur interaction avec leur cible moléculaire (ARN et protéine). En plus de leur pouvoir antibiotique, les aminoglycosides sont également des ligands universels pour des ARN, capables d’interagir spécifiquement avec notamment les ARN du VIH-1 suivants : DIS, TAR, RRE. L’élaboration d’aminoglycosides modifiés présente un énorme avantage car le domaine d’application, et en conséquence les retombées, sont grandes. Néanmoins, la complexité structurale de ces molécules est un frein majeur, la fonctionnalisation chimiosélective est indispensable mais malheureusement peu décrite dans la littérature. Dans le cadre de ce travail, nous avons développé deux types d’approches pour cibler le DIS et/ou le site A du ribosome bactérien. La première originale, mais risquée se base sur le concept de click in situ. La seconde approche est traditionnelle et est basée sur la fonctionnalisation sélective de certaines positions clés des aminoglycosides. / The development of new antibiotics is a major public health issue. Given the high potential of aminoglycosides as antibiotics, these compounds have aroused great interest in many research groups. However, despite their maturity, their use is still limited because of their toxicity and the increasing development of resistance mechanisms to aminoglycosides. To better understand the problems inherent to their use, it is crucial to understand their action a cellular level, and to study the interactions with their molecular targets (RNA and protein). In addition to their antibiotic power, aminoglycosides are also universal ligands for several RNAs, capable of specific interactions with RNAs of HIV-1: DIS, TAR and RRE. The elaboration of modified aminoglycosides presents a huge advantage because the domain of application, and therefore the benefits, are important. Nevertheless, the structural complexity of these molecules is a major constraint, chemoselective functionalization is essential but unfortunately poorly described in the literature.In this work, we developed two approaches to target the DIS and/or the A site of the bacterialribosome. The first one, unique but challenging is based on the concept of in situ click chemistry. The second approach is conventional and is based on the selective functionalization of some keypositions of aminoglycosides.

Substituted tetrazines have been found to undergo facile inversed electron demand Diels-Alder reactions with “tunable” reaction rates. By varying the substituents on tetrazine, cycloaddition rate variations of over 200 fold have been achieved with the same dienophile. Coupled with the availability of different dienophiles, such as norbornene, the reaction rate difference can be over 14,000 folds. These substituted tetrazines can be very useful for selective labeling under different conditions. This finding paves the way to utilize tetrazine conjugation reactions for not only DNA but also stage labeling work. Carbon monoxide (CO) belongs to the gasotransmitter family of signalling molecules in the mammalian systems with importance on par with that of NO and H2S. Studies have shown that endogenous production of CO has anti-inflammatory, anti-proliferative, and anti-apoptotic effects in mammalian system. Besides of the conventional metal-based carbon monoxide releasing molecules (CORMs) to deliver CO for therapeutic purposes, organic CO prodrugs represent a new direction. Here we report the “click and release” approached to release CO. Unlike the metal-based CORMs, our system does not contain transition metal and liberates CO with controllable manner and possesses potential tunable releasing rate property under physiological conditions.

Les glioblastomes sont des tumeurs cérébrales qui sont extrêmement agressives, et qui, en dépit de l'arsenal thérapeutique (chirurgie, radiothérapie ou chimiothérapie), ne laissent pas plus de 16 mois d'espérance de vie aux patients. Dans le cadre de cette thèse, nous proposons d'utiliser certaines toxines en tant que vecteurs pour l'administration de médicaments anticancéreux, et notamment pour le traitement du gliome. Les travaux présentés ici se concentrent sur l’utilisation de variants de la maurocalcine (MCa) et des analogues de la chlorotoxine (CTX). La MCa est une toxine issue du venin du scorpion Scorpio maurus palmatus, qui est capable de pénétrer dans les cellules facilement et rapidement. Il a été prouvé que la MCa peut entrer dans la cellule avec une cargaison. C’est en exploitant cette capacité présente chez deux de ces variants que nous avons synthétisé avec succès deux nouveaux composés à base de cette toxine avec de la doxorubicine et un dérivé du platine. Les études de toxicité et de caractérisation de ces composés qui ont été réalisé on permit de mettre en évidence l’intérêt et le potentiel de la MCa. La seconde partie de ces travaux de thèse portée sur la CTX et des peptides semblables, également extrait de venin de scorpion. Ils ont la particularité de fixer / interagir uniquement avec les cellules cancéreuses d'origine gliale. Après une rapide caractérisation de ces analogues de la CTX, l’un d’eux la Lqh-8/6 a été utilisé avec succès pour l'administration ciblée de doxorubicine. L’ensemble des travaux menés durant cette thèse constitue une base de départ solide pour une amélioration des systèmes de vectorisation, surtout en cancérologie de molécules actives. De plus ces résultats mettent aussi en avant l’avantage de l’utilisation d’un système de couplage « universel » basé sur la chimie click. / Glioblastoma are cerebral tumors that are extremely aggressive, and that, in spite of a battery of therapeutic interventions (surgery, radiotherapy or chemotherapy), leave no more than 16 months life expectancy to the patients. As part of this thesis, we propose to use some selected toxins as vectors for the delivery of anticancer drugs, and namely for the treatment of glioma. The works presented here concentrate on the use of variants of maurocalcine (MCa) and the analogues of chlorotoxine (CTX). MCa is a toxin from of the scorpion Maurus palmatus that has cell penetrating propriety. It has been proved that MCa can enter the cell with cargoes. While exploiting this present capacity to two of these variants we synthesized successfully two new compounds with this toxin with the doxorubicine and a by-product of the platinum. Toxicity studies and characterization of these compounds that have been made were permitted to highlight the interest and potential of the MCa. The second part of the thesis work focused on the CTX and similar peptides, also extracted from scorpion venom. They have the particularity to fix/ interact only with cancer cell from neuroectodermal origin. After a fast characterization of these analogues of CTX, one of them (Lqh-8/6) was successfully used for the targeted administration of doxorubicin. All work conducted during this thesis constitutes a solid starting point for an improvement of the systems of vectorization of active molecules, especially in cancer research Moreover, these results also emphasize the advantage of the use of a system of "universal" coupling based on the click chemistry.

Des nanocapsules (NCs) biodégradables contenant une substance active et destinées à des applications environnementales ont été élaborées par un procédé d’émulsion-évaporation de solvant couplé à une réaction de chimie « click » interfaciale. Deux types de réactions « click » ont été testés: (i) cycloaddition azide-alcyne catalysée par le Cu(I) et (ii) thiol-ène. Ces NCs sont constituées d’une écorce en polymère hydrophobe (polylactide) entourant un cœur liquide (Miglyol®810) et recouverte d’une couronne hydrophile polysaccharide (dextrane). Des nanosphères (sans cœur liquide) ont aussi été produites. Ces nano-objets ont été caractérisés en termes de distribution de tailles, morphologie, taux et épaisseur de recouvrement en dextrane ainsi qu’efficacité de couplage « click ». La stabilité colloïdale en milieu salin et la stabilité du recouvrement en présence d’un tensioactif compétitif ont été étudiées. Enfin, une substance active a été encapsulée et libérée à partir des nano-objets / Biodegradable nanocapsules allowing encapsulation of active substances for environmental applications were produced by emulsion-evaporation method combined with a “click” reaction occurring at the liquid/liquid interface of emulsion droplets. Two types of “click” reaction were tested: (i) copper-catalyzed azide-alkyne cycloaddition (CuAAC) and (ii) thiol-ene reaction. The NCs are composed of a hydrophobic polymer shell (polylactide), a liquid core (Miglyol®810) and a hydrophilic polysaccharide coating (dextran). For comparison, nanospheres (without oily core) were also prepared. These nano-objects were characterized in terms of size distribution, dextran coverage density and thickness, “click” coupling efficiency and morphology. Colloidal stability in NaCl solutions as well as dextran coverage stability against an anionic competitive surfactant were also studied. Finally, an active substance was encapsulated and released from these nano-objects

Os Clusters de acetato de rutênio de fórmula geral [Ru3O(AcO)6L3]n proporcionam blocos de montagem interessantes para sistemas supramoleculares, exibindo características eletrocrômicas e redox reversível, cobrindo uma ampla faixa de potenciais em solventes aquosos e orgânicos. Nesta Tese, estas espécies foram combinadas com o ligante tridentado, 4\'-piridil-2,2\':6\',2\"-terpiridina (pytpy), produzindo complexos do tipo [Ru3O(AcO)6(pytpy)3]n e [Ru3O(AcO)6(py)2(pytpy)]n, que foram totalmente caracterizados em estado sólido e solução. Nestes complexos a terpiridina permanece disponível para ligar-se a íons metálicos como o Fe(II), gerando um complexo binuclear de baixo spin muito estável, do tipo {Fe[Ru3O(AcO)6(py)2(pytpy)]2}n. Este processo pode ser monitorado eletroquimicamente, partindo de íons Fe(III), que exibem uma baixa afinidade pela terpiridina central, e ciclando o potencial para gerar íons de Fe(II). Desta forma, pode-se executar um procedimento típico de click chemistry, que leva a uma estrutura estendida, Fe-cluster-pytpy na superfície do eletrodo. Este filme conserva a funcionalidade dos blocos de montagem, permitindo aplicações interessantes em dispositivos moleculares, tais como sensores e smart Windows. / Ruthenium acetate clusters of general formula [Ru3O(AcO)6L3]n provide interesting building blocks for assembling supramolecular systems, displaying reversible redox and electrochromic characteristics, over a wide range of potentials in aqueous and organic solvents. In this thesis, such species has been combined with the tritopic, 4\'-pyridil-2,2\':6\',2\"-terpiridine (pytpy) ligand, yielding complexes of the type [Ru3O(AcO)6(pytpy)3]n and [Ru3O(AcO)6(py)2(pytpy)]n, which have been fully characterized in solid state and solution. In these complexes, the terpyridine moiety remains available for binding metals ions, such as Fe(II), generating very stable, low spin binuclear complexes, of the type n. This process can be monitored electrochemically, starting from Fe(III) ions, which exhibits a much lower affinity for the terpyridine center, and cycling the potentials in order to generate Fe(II) ions. In this way, one can start a click chemistry, ending up with an extended structure of Fe-cluster-pytpy film at the electrode surface. This film preserves the functionality of the building block complexes, a llowing many possible applications in molecular devices, such as sensors and smart windows.

Neste trabalho promovemos a síntese de sulfóxidos vinílicos ?-substituídos através da reação de acoplamento cruzado de Suzuki-Miyaura. Também foi feita a síntese de sulfóxidos enínicos inéditos, pela adição do nucleófilo no carbono β-sulfóxido. Esses compostos eram passíveis de serem submetidos a reação de rearranjo de Pummerer aditivo e assim gerarem uma pequena biblioteca de compostos α-tioaldeídos. Um desses aldeídos sintetizados foi empregado na reação de formação de uma imina propargílica, com consequente reação de CuAAC formando iminas triazólicas. Outras iminas arílicas foram sintetizadas, passando por uma etapa de redução, com intuito de se obter a amina livre, para que fosse feita a reação de ciclização com auxílio de um agente eletrofílico. Outra classe de composto organoenxofre foi sintetizada, as N-sulfinil imina, que após a reação de acoplamento cruzado de Sonogashira, com consequente remoção de um grupo protetor e a formação do anel heterocíclico, foram obtidos compostos triazólicos N-sulfinil imínicos. / In this work we promote the synthesis of α-substituted vinylic sulfoxides through the Suzuki-Miyaura cross coupling reaction. Also the synthesis of unpublished enynic sulfoxides was made, by the addition of the nucleophile in the β-sulfoxide carbon. These compounds were susceptible to additive Pummerer rearragement reaction and thus generated a small library of compounds. One of these aldehydes synthesized was used in the formation reaction of a propargyl imine, with consequent CuAAC reaction, forming triazol imines. Other aryl imines were synthesized, undergoing a reduction step, in order to obtain the free amine, so that the cyclization reaction was carried out with the aid of an electrophilic agent. Another class of organosulfur compound was synthesized, the N-sulfinyl imine, which after the Sonogashira cross-coupling reaction, with consequent removal of a protecting group and formation of the heterocyclic ring, N-sulfinyl imine triazolic compounds were obtained.

Parmi les composants de la biomasse, la lignine est considérée comme l'un des plus prometteurs polymères naturels qui convient à la conversion de la biomasse en valable produits chimiques et matériaux. Malgré une grande quantité de lignine est générée dans l'industrie papetière, seule 2% est exploitée dans l'industrie chimique. La présence de soufre et la grande diversité moléculaire sont les principaux obstacles pour l'utilisation de la lignine. La modification chimique a été reconnue comme un outil pour contourner ces limites. Dans cette thèse, différentes stratégies de synthèse ont été appliquées pour introduire de nouveaux groupes chimiques sur une soude lignine que présents une haute fonctionnalisation de groups hydroxyles. Les dérivés correspondants de lignine ont été utilisés soit pour l’élaboration des matériaux par click polymérisations, soit pour augmenter l’hydrophobicité de la lignine à la fine de faciliter son traitement avec des matrices polymériques. / Among biomass components, lignin is considered one of the most promising natural polymers suitable for the conversion of biomass into renewable added-value chemicals and materials. However, large amount of lignin generated from wood pulping industry is burn as low cost energy source, and only 2% is exploited in the chemical industry. The presence of sulphur moieties and the large molecular diversity are the most reasons impeding the use of lignin as building blocks for the production of chemicals and materials. Chemical modifications have been acknowledged to be an important tool to circumvent these limitations. In the current work, taking advantage of the high hydroxyl groups content of a sulphur free soda lignin (SL), different synthetic strategies have been applied to introduce new chemical groups and used either to produce lignin derivatives suitable for “click” polymerization either to increase lignin hydrophobicity, facilitating its processing in polymeric matrices.

The aim of this research was to find relationships between complimentary advertising strategies and sponsored search advertisement (SSA) in order to formulate a model to maximise return on investment achieved from online sponsored search advertisements. The results obtained from statistical analyses of SSA campaign data showed that complimentary online and offline advertisement campaigns have various different correlations to impressions, click-through rates, number of pages visited, time spent visiting a website, bounce rate of visitors to the website, cost-per-click and number of new registrations per keyword search from visitors gained through SSA campaigns. In particular, online display advertisements were found to have a slight positive correlation with new registrations made by customers gained through a simultaneously running SSA campaign. Offline radio adverts were found to have a positive correlation with impressions gained for SSA campaigns, whilst at the same time showing a negative correlation with the number of pages viewed by website visitors obtained through the SSA campaign. Some negative correlations to SSA campaign performance were also found, with the time visitors spent viewing the website decreasing, their bounce rate increasing and the cost-per-clicks for the keywords in the SSA campaign also increasing during periods when offline radio adverts were active. Offline television adverts were found to have a negative correlation with impressions gained for SSA campaigns, as well as the click-through rate for the keywords in these SSA campaigns. Offline television adverts did however also show a negative correlation with the cost-per-clicks for keywords in the SSA campaigns. Finally, a graphical model was developed to illustrate these correlations found between complimentary advertisement campaigns and SSA performance metrics. / Dissertation (MBA)--University of Pretoria, 2012. / Gordon Institute of Business Science (GIBS) / unrestricted

Les anticorps monoclonaux anti-tumoraux sont produits en cellules eucaryotes. Pour des raisons de temps et de cout, peu de candidats sont sélectionnés après des tests in vitro pour être produits à large échelle et testés in vivo. Pour tester plus d’anticorps plus rapidement, nous souhaitons produire des fragments variables simple chaine (scFv) chez E. coli. et les coupler à un fragment constant Fc par chimie click pour reconstituer des mimes d’immunoglobulines naturelles. Cette production indépendante des fragments est aussi un outil modulaire permettant de combiner rapidement un grand nombre scFv et de Fc différents.La chimie click est basée sur une réaction spécifique et de haut rendement entre un azide et un cyclooctyne. Les fragments ont donc été fonctionnalisés sur des résidus spécifiques (tags) par des composés chimiques pour introduire ces fonctions à leur extrémité. La première étape a consisté à introduire des tags en C-terminal du scFv anti-HER2 4D5 et en N-terminal du Fc d’IgG1 humaine. Les scFv ont été produits en cytoplasme d’E. coli à hauteur d’au moins 100 mg/L puis oxydés in vitro au sulfate de cuivre. Le fragment Fc a été produit classiquement en cellules humaines. Cinq réactions chimiques ou enzymatiques ont été optimisées et comparées en termes de spécificité et de rendement. La conjugaison d’une amine sur un tag glutamine catalysée par la transglutaminase microbienne a donné les meilleurs résultats. Le scFv a ainsi été dérivé par l’azadibenzocyclooctyne et le Fc par l’azide à hauteur de 60-70%. Lorsqu’ils sont mélangés, ces fragments forment un (scFv)2-Fc et un scFv-Fc avec des rendements globals respectifs de 10-20% et 20-30% après optimisation.Les mélanges scFv + Fc après réaction de chimie click se fixent de la même façon que le (scFv)2-Fc eucaryote au récepteur HER2. Il reste désormais à montrer que leur capacité à inhiber la prolifération d’une lignée exprimant ce récepteur est similaire. L’objectif final est d’obtenir une inhibition de croissance tumorale similaire sur des xénogreffes. / Anti-tumoral monoclonal antibodies are currently produced in eukaryotic cells. For cost and time reasons, a limited number of potential candidates are selected after in vitro tests. They are produced at large scale and then tested in vivo. To test more antibodies and more rapidly, we chose to produce single chain variable fragments (scFv) in bacteria, and to couple them to the eukaryotic constant fragment (Fc) thanks to click chemistry to reconstitute immunoglobulin-like compounds. For a given cost, this enables to produce and test in vivo a larger number of clones. This independent production of fragments is also a flexible tool allowing the combination of different Fc isotypes/allotypes with different scFvs.Click chemistry is based on a specific and high-yield reaction between and azide and a cyclooctyne. Therefore, antibody fragments were functionalised on specific residues (tags) by chemical linker so that each part will contain one of these chemical moieties at their extremity. The first step consisted in introducing tags into the anti-HER2 scFv 4D5 C-terminus and human IgG1 Fc N-termini sequences. The scFvs were produced with yields higher than 100 mg/L in the E. coli cytoplasm and in vitro oxidized with copper sulfate. The Fc fragment was classically produced in human cells. Five chemical or enzymatical reactions were optimised and compared in terms of specificity and yield. The coupling between an amine and a glutamine tag catalysed by microbial transglutaminase gave the best results. The scFv fragment was thus functionalised with an azadibenzocyclooctyne and the Fc fragment with an azide at 60-70%. When mixed together, these fragments formed a (scFv)2-Fc and a scFv-Fc with global yields respectively of 10-20% and 20-30% after optimisation.After the click reaction, the scFv + Fc mix binds to the HER2 receptor on the same way as the eukaryotic (scFv)2-Fc in terms of HER2-binding and proliferation inhibition capacity. Now, it must be demonstrated that their proliferation inhibition of a HER2-positive cell line is similar. The final aim is to get a similar tumour growth inhibition on murine xenografts.