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Thesis (M.Sc.). / Written for the Institute of Parasitology. Title from title page of PDF (viewed 2008/07/29). Includes bibliographical references.

Thesis (M.Sc.). / Written for the Institute of Parasitology. Title from title page of PDF (viewed 2008/07/29). Includes bibliographical references.

Current FDA approved anti-cytomegalovirus antivirals are ganciclovir, foscarnet, and cidofovir. These drugs target the viral polymerase to inhibit DNA synthesis. Halogenated benzimidazoles target a step later in viral replication during packaging and cleavage of the viral genome. The compounds 2-Bromo-5,6-dichloro-1-(β-D-ribofuranosyl)benzimidazole (BDCRB) and 2,5,6-trichloro-1-(β-D-ribofuranosyl)benzimidazole (TCRB) are novel inhibitors of human cytomegalovirus (HCMV) and resistance to these compounds are found within the human cytomegalovirus viral terminase. Terminase is unique to the virus and in theory provides a good target for antiviral development. Beginning with a BDCRB resistant guinea pig cytomegalovirus observations found a single mutation located in the viral terminase gene UL89 and unique formation found in genomic ends. In guinea pigs, the virus continued to produce large amounts of monomer. This is in contrast to human cytomegalovirus treated with either BDCRB or TCRB. Both compounds produced very little monomer DNA but created a supergenomic species, called monomer-plus that migrates to the apparent size of 270-kb on a pulse field gel. A model was developed to explain formation of monomer plus and my project aims originated from the model. In the presence of BDCRB and TCRB, packaging is relaxed resulting in the normal cleavage site being skipped and cleavage occurring at the next available cleavage site creating a short-long-short genome. In guinea pigs, cleavage has been relaxed by premature cleavage of the terminal ends. Current antivirals do not block viral entry therefore, patients are infected with HCMV. Blocking entry of HCMV into cells would prevent HCMV infection. It has been shown that antibodies to epitopes within the gH/gL/UL128-131 complex can block viral entry into endothelial, epithelial, and other cell types while antibodies to gB block entry into fibroblasts. The majority of the current work on entry into epithelial cells has been performed in retinal pigment epithelium and epithelial cells derived from tumors. Viral entry into cell lines derived from mucosa cells was dependent on the complex gH/gL/UL128-131. Rabbit antibodies raised against UL130 and UL131 peptides neutralized epithelial entry with effects as potent as human seropositive sera. This suggests that the entry complex can be blocked by single epitopes.

Ce manuscrit décrit la synthèse et la conception de 3 nouvelles familles de molécules, les [alpha],[alpha]-diméthylamines, les 6-nitroaminopyridines et les benzimidazoles substitués. L’étude des [alpha],[alpha]-diméthylamines nous a amené à développer une voie de synthèse originale en utilisant les propriétés chimiques du chlorure de cérium, qui, lorsqu’il est complexé avec un organolithien permet d’alkyler deux fois une fonction nitrile. Une étude exhaustive des propriétés chimiques du cérium et du chlorure de cérium en synthèse organique a également été menée. La synthèse totale de la 6-nitro-2-aminopyridine nous a amené à développer une voie de synthèse générale permettant de conduire aux différents isomères de position de ce composé. Cette voie a également été optimisée et transposée à une unité de synthèse de 50L. Les différentes conditions réactionnelles de cyclisation de diamines fonctionnalisées nous ont permis de synthétiser des séries de 5 benzimidazoles fonctionnalisés : les benzimidazoles, les 2-méthylbenzimidazoles, les benzimidazolones, les 2-chlorobenzimidazoles et les 2-mercaptobenzimidazoles. Une revue exhaustive des différentes méthodes de préparation de ces benzimidazoles est également détaillée et ont servi de base à ce travail de synthèse. / The present report describes the design and the synthesis of 3 new family of molecules, the [alpha],[alpha]-dimethylamines, the 6-nitroaminopyridines and substituted benzimidazoles. The study of [alpha],[alpha]-dimethylamines devellop an original synthetic route using the chemical properties of cerium chloride, witch can alkyl two times a nitril fonction when it is complexed with organolithium reagent. Also, an exhaustive study was carried on chemical properties of cerium and cerium chloride in organic synthesis. The total synthesis of 6-nitro-2-aminopyridine bring us to devellop a general synthetic route leading to different position isomeres of this compound. Also, this route was optimized and transposed to a 50L synthesis unity. The different conditions of cyclisation of substituted diamines allowed us to synthetise 5 series of fonctionnalized benzimidazoles : the benzimidazoles, the 2-methylbenzimidazoles, the benzimidazolones, the 2-chlorobenzimidazoles and the 2-mercaptobenzimidazoles. An exhaustive review of the different methods for preparing benzimidazoles was carried and was the basis for this synthetic work.

Foram estudados 83 pacientes com esofagite erosiva graus I e II, pela classificação de Savary-Miller modificada, divididos em 3 grupos. Um sem Helicobacter pylori, dois outros com Helicobacter pylori, com e sem o gene cagA. Avaliou-se a participação da bactéria e de seu gene cagA, associados à estudo histopatológico de antro e corpo e à gastrinemia basal, na cicatrização da mucosa do esôfago após tratamento com pantoprazol 40 mg ao dia por 6 semanas. Verificou-se que a presença do Helicobacter pylori, independentemente da presença do gene cagA, facilita a cicatrização esofágica. Indivíduos com gastrinemias maiores também tendem a cicatrizar melhor. Não houve relação do resultado do estudo histopatológico com a resposta terapêutica / Eighty three patients with grade I-II of the modified Savary-Miller classification have been studied. They were divided in three groups. One without Helicobacter pylori infection, two with the bacterium, one with and other without the cagA gene. We verified the influence of cagA status, histopathology of antrum and body of the stomach and gastrinemia in the esophageal healing rates after treatment with pantoprazole 40 mg once a day for six weeks. Helicobacter pylori presence but not cagA status and gastrinemia led to better healing rates. Histopathology of the gastric mucosa did not influence the response

Preclinical methods for the identification and characterization of molecules for development into new cancer drugs were investigated. Based on repurposing, i.e. the exploration of currently prescribed drugs for new indications, and as a result of a new high throughput screening (HTS) approach, the benzimidazoles benomyl and carbendazim, the alkaloid cryptolepine and the acridine acriflavine were found interesting to characterize using these methods. In mice the benzimidazoles inhibited 3H-thymidine incorporation in tissues with high cell renewal, with benomyl being more active than carbendazim.  They were rapidly absorbed with highest amounts seen in the liver, kidneys and gastro-intestinal lumen as evidenced from distribution of 14C-labeled drugs. In human tumour cell lines, the benzimidazoles showed a similar activity pattern but benomyl was more potent. This was true also in tumour cells from patients but carbendazim was relatively more active against solid tumours. Analyses of drug activity cross-resistance patterns and of drug activity – gene expression correlations in a cell line panel suggested multiple mechanisms of action for the benzimidazoles. Cryptolepine was widely distributed to tissues in vivo in the mice. It was more potent than the benzimidazoles in tumour cells, with highest activity in haematological malignancies but some patient samples of breast, colon and non small-cell lung cancer were sensitive. Cross-resistance analysis indicated cryptolepine to be a topoisomerase II inhibitor whereas drug activity – gene expression correlations suggested additional mechanisms of action. HTS on 2 000 molecules in colon cancer cell lines and normal cells identified acriflavine as a hit molecule, subsequently shown to have unprecedented activity against colorectal cancer tumour cells in patient tumour samples. Connectivity map analysis, based on drug induced gene expression perturbation patterns in a tumour cell line, indicated acriflavine to be a topoisomerase inhibitor, subsequently confirmed in a plasmid relaxation assay. In conclusion, repurposing of drugs and HTS using stringent activity criteria followed by preclinical characterization might contribute to more efficient development of new cancer drugs.

Made available in DSpace on 2014-10-09T12:53:32Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T13:58:56Z (GMT). No. of bitstreams: 0 / Dissertação (Mestrado) / IPEN/D / Instituto de Pesquisas Energéticas e Nucleares - IPEN-CNEN/SP

Tridentate benzimidazole-based ligands, bis((1H-benzimidazol-2-yl)methyl)sulfide (BNSN) and bis((1H-benzimidazol-2-yl)methyl)amine (BNNN), along with dinonylnaphthalene sulfonic acid (DNNSA) as a synergist, were investigated as potential selective extractants for Ni2+ from base metals in a solvent extraction system using 2-octanol/Shellsol 2325 (8:2) as diluent and modifier. However, extraction studies show a lack of pH-metric separation of the later 3d metal ions with bis((1-octylbenzimidazol-2-yl)methyl)sulfide (BONSN) and bis((1- decylbenzimidazol-2-yl)methyl)amine (BDNNN) as extractants, but extractions occurred in the low pH range with an opportunity for back extraction. This investigation suggested that tridentate ligands (at least those of the nature investigated here) are not feasible extractants for separation of base metal ions due to their lack of stereochemical “tailor-making.”

Made available in DSpace on 2014-10-09T12:53:32Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T13:58:56Z (GMT). No. of bitstreams: 0 / O presente trabalho apresenta os dados obtidos no estudo para o desenvolvimento de método multirresíduo de pesticidas dos grupos de benzimidazol (carbendazim, tiabendazol e tiofanato metílico), carbamatos (aldicarbe, aldicarbe sulfona, aldicarbe sulfóxido, carbaril, carbofurano, metomil, metiocarbe, pirimicarbe, propoxur) e triazinas (atrazina e simazina) em amostras de milho verde. Os pesticidas foram extraídos em acetona, sob agitação, diluídos em água e injetados no sistema LC/MS/MS. A técnica analítica de quantificação e confirmação utilizada foi a Cromatografia Líquida acoplada à Espectrometria de Massas em Tandem com ionização por Electrospray no modo positivo. A validação do método multirresíduo foi submetida em conformidade com a norma EC/2002/657 da Comunidade Européia. Para quantificação dos analitos utillizouse as curvas analíticas dos princípios ativos, nas concentrações que variaram de 0,04 a 8,0 ng/ml, correspondentes a 2,0 a 400 ?g.kg-1 na amostra. Para o estudo da recuperação, os pesticidas foram avaliados em cinco níveis, de ½ Limite de quantificação (LOQ) a 5 LOQ, correspondentes a níveis de fortificação de 4,0 a 200 ?g.kg-1 . Os resultados da recuperação apresentaram, em níveis aceitáveis, na faixa de 80 a 110% e com precisão satisfatórias, CV <=20%, com exceção de aldicarbe e de aldicarbe sulfona. Os limites de quantificação do método variaram de 8 a 40 ?g.kg-1 e os limites de detecção do método, de 0,2 a 2,9 ?g.kg-1. Os limites de quantificação do método atendem aos limites máximos de resíduos da legislação brasileira em vigor. Nas amostras estudadas, não foram encontrados resíduos de pesticidas acima do limite de quantificação do método. Por outro lado, todos os pesticidas, exceto carbaril e pirimicarbe, foram detectados em todas as amostras e em níveis acima dos limites de detecção do método. A alta incidência da presença de resíduos de pesticidas se deve possivelmente ao uso inadequado de agrotóxico, quando não autorizado o seu uso para a cultura de milho no Brasil ou proveniente de alguma contaminação, como o tratamento de agrotóxicos aplicados em outras culturas plantadas no mesmo solo. / Dissertação (Mestrado) / IPEN/D / Instituto de Pesquisas Energéticas e Nucleares - IPEN-CNEN/SP