Orientador: Renato Henriques de Souza / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Química / Made available in DSpace on 2018-08-18T09:37:25Z (GMT). No. of bitstreams: 1 Souza_RenatoHenriquesde_D.pdf: 4255462 bytes, checksum: d86f248ffcc38a9903cedbcccb4265a4 (MD5) Previous issue date: 2011 / Resumo: Anéis de três membros são considerados excelentes reagentes de partida para a síntese orgânica. Dentre estes, destacamos as diaziridinonas, que são heterociclos de três membros contendo dois átomos de nitrogênio e um grupo funcional carbonílico exocíclico, e comportam-se como uma classe especial destes heterociclos. Diaziridinonas que possuem substituintes volumosos apresentam reatividade muito baixa frente à nucleófilos, ao contrário do comportamento da maioria dos heterociclos de três membros. No entanto, em diversas reações envolvendo ataque nucleofílico ao carbono carbonílico da 1,2-di-t-butildiaziridinona o CoCl2 tem um papel marcante, ativando o heterociclo e permitindo o ataque, levando à formação de compostos polifuncionalizados. Explorando este potencial sintético, resolvemos estudar reações entre diaziridinonas e nucleófilos orgânicos bifuncionalizados, na presença e ausência do catalisador. Em reações com aminoálcoois, catalisadas, e efetuadas na proporção estequiométrica 1,2-di-t-butildiaziridinona: nucleófilo de 1:1, obtivemos novas rotas sintéticas para importantes produtos heterocíclicos, que são as 1,3-oxazolidin-2-onas e as 1,3-oxazinan-2-onas. Em reações análogas efetuadas na proporção 2:1 obtivemos uma diferente classe de produtos, inéditos, e de cadeia aberta. Não obtivemos produtos nas reações realizadas sem CoCl2 e não isolamos produtos ao utilizarmos a 1,2-di-t-octildiaziridinona, mais impedida estericamente. Em reações da 1,2-di-t-butildiaziridinona com aminoácidos obtivemos apenas produtos de cadeia aberta, resultantes do ataque nucleofílico do grupo amino ao carbono carbonílico do heterociclo, e estas reações são favorecidas na presença de CoCl2. Em alguns casos isolamos também 1,3-di-t-butiluréia, resultante da redução do heterociclo. Nas reações entre 1,2-di-t-butildiaziridinona e hidroxiácidos isolamos produtos resultantes do ataque nucleofílico do grupo ácido ao carbono carbonílico do heterociclo, levando a produtos de cadeia aberta. Estas reações são desfavorecidas na presença de CoCl2, e os produtos formados são os mesmos, independentemente da proporção estequiométrica dos reagentes. Em comum, todos os exemplos mostram o elevado potencial sintético da 1,2-di-t-butildiaziridinona na obtenção de diversas classes de produtos polifuncionalizados / Abstract: Three-membered rings are considered to be excellent starting reagents for organic synthesis. Among these, we feature the diaziridinones, which are three-membered heterocycles containing two nitrogen atoms and an exocyclic carbonyl functional group, and they behave as a special class of those heterocycles. Diaziridinones possessing bulky substituents have very low reactivity towards nucleophiles, unlike the behavior of most three membered heterocycles. However, in several reactions involving the nucleophilic attack to the carbonyl carbon atom of 1,2-di-t-butyldiaziridinone, CoCl2 has a remarkable role, activating the heterocycle and allowing the attack, leading to polyfunctional compounds. Exploring this synthetic potential, we studied reactions between diaziridinones and bifunctionalized organic nucleophiles in presence and absence of the catalyst. In reactions with aminoalcohols, catalyzed, and performed at the stoichiometric 1,2-di-t-butyldiaziridinone: nucleophile ratio of 1:1, we obtained new synthetic routes for important heterocycles, which are the 1,3-oxazolidin-2-ones and 1,3-oxazinan-2- ones. In similar reactions performed in a 2:1 ratio we obtained a different class of up to date unpublished open-chain products. We did not obtain products in reactions performed without CoCl2 and we did not isolate any product when we used more sterically hindered 1,2-di-t-octildiaziridinone. In reactions of 1,2-di-t-butyldiaziridinone with amino acids we obtained only open-chain products, resulting from nucleophilic attack of the amino group to carbonyl carbon of the heterocycle, and these reactions are favored in the presence of CoCl2. In some cases we also isolated 1,3-di-t-butilurea, resulting from the reduction of the heterocycle. In the reactions between 1,2-di-t-butyldiaziridinone and hydroxy acids we isolated products resulting from nucleophilic attack of the acid group to the carbonyl carbon of the heterocycle, leading to open-chain products. These reactions are unfavored in the presence of CoCl2, and the products formed are the same, regardless of the stoichiometric amounts of reagents. In common, all examples show the high synthetic potential of 1,2-di-t-butyldiaziridinone as starting material for diverses classes of polyfunctional products / Doutorado / Quimica Inorganica / Doutor em Ciências

The inner surface of the stomach is lined by a mucous membrane known as the gastric mucosa. The integrity of the gastric mucosa is critical for protecting the stomach from the low pH and proteolytic environment within the lumen. Both clinically and experimentally, exposure of gastric mucosal cells to bile salts is known to cause injury. Bile salts present in duodenogastric reflux are thought to play a significant role in gastric ulcer formation and alkaline gastritis. In vitro, studies using physiologic concentrations of the secondary bile salt, deoxycholic acid, indicate that bile salts can induce apoptosis in cultured human gastric epithelial cells in a caspase-dependent manner. Therefore, there is interest in developing approaches that can protect gastric cells from bile salt-induced damage. It has been shown that induction of the stress protein, heme oxygenase-1, can provide protection against apoptosis. Therefore, the objective of this study was to test the hypotheses that heme oxygenase-1 expression could be induced in human gastric epithelial cells and that furthermore; this would provide protection from deoxycholic acid-induced apoptosis. Heme oxygenase-1 expression was induced pharmacologically or by introduction of a plasmid expressing heme oxygenase-1 into the gastric epithelial cell line, AGS. Induction of heme oxygenase-1 prior to challenge with deoxycholate reduced apoptotic-associated morphological changes, DNA fragmentation, the appearance of oligonucleosomes in the cytoplasm, and activation of caspase-3 and caspase-9. Based on these results, it was concluded that expression of heme oxygenase-1, or the introduction of its products, can provide protection to human gastric epithelial cells against sodium deoxycholic acid induced-apoptosis.

A systems biology approach was adopted in order to assess various aspects of the disease oral squamous cell carcinoma. Three main aims were addressed; assess the ability of CoCl2 to mimic the hypoxic response in a eukaryotic cell line, assess the role of PDE4D in oral squamous cell carcinoma (OSCC) and the construction of a normoxic/hypoxic mathematical model to identify therapeutic targets.Cancer cells often acquire a revised metabolism which aids in initiation, survival and progression of the tumour. This is predominantly due to the transcription factor HIF-1 which is activated under hypoxic conditions. Certain compounds such as cobalt chloride (CoCl2) have been used extensively to inhibit the degradation of HIF-1α and simulate hypoxia. CoCl2 is likely to have off-target effects on metabolism; these effects were examined when exposing human telomerase reverse transcriptase (hTERT) cells to 100μM CoCl2. Gas chromatography-mass spectrometry (GC-MS), liquid chromatography-mass spectrometry (LC-MS) based metabolomics were utilised in combination with ELISA assays for HIF-1α and ATP. Central metabolism was accurately mimicked when hTERT cells were subjected to 100μM CoCl2, however; it was apparent that this concentration of CoCl2 does not induce an equal extent of hypoxia as 1% oxygen. A number of off-target effects of CoCl2 were observed in secondary metabolism, specifically in lipids and fatty acids. In conclusion, CoCl2 should be used with caution as a hypoxic mimicker with the caveat that interpretation of results should be restricted to its effects on central metabolism.The transcription factor CREB has the ability to regulate approximately 4000 genes, a number of which are associated with cancer initiation and progression. Cyclic adenosine monophosphate (cAMP) is required to activate CREB and is partially regulated through its degradation via the enzyme phosphodiesterase type 4D (PDE4D). A homozygous deletion of PDE4D has been associated with OSCC; however; the exact consequence of this deletion has not been fully elucidated. PDE4D was knocked down in the OSCC cell line BicR16 and cellular proliferation, migration, resistance to ionising radiation and central metabolism was investigated using MTT, scratch, clonogenic and GC-MS, respectively. The knockdown resulted in an increase in proliferation, migration and radiation resistance suggesting the role of PDE4D as a TSG. Amino acids, cholesterol, fatty acids, carbohydrates and TCA intermediates were found to be altered in concentration.A mathematical model of glycolysis, TCA and glutaminolysis under normoxia and hypoxia was constructed through the amalgamation of two established models from the literature. New reactions, parameters and metabolite concentrations were added and unnecessary entities were deleted. COmplex PAthway SImulator (COPASI) was utilised to construct the model before validating the model using experimental data from the literature and steady state and flux analyses. Sensitivity analysis and a reduction in external glucose and glutamine were mimicked and the alterations in hypoxic and normoxic metabolism analysed. The reactions vCSII, vGS, vPGK and vGII were identified as potential therapeutic targets which may affect metabolism in hypoxia only. However, certain validation methods proved unsuccessful and hence the model requires further work before attempting the analyses again.