Submitted by Irene Nascimento (irene.kessia@ufpe.br) on 2017-03-24T17:41:49Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Dissertação Mestrado - Tiago Barros Santos Malta.pdf: 8207831 bytes, checksum: a5e3ee5357f988c47072c9481326377e (MD5) / Made available in DSpace on 2017-03-24T17:41:49Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Dissertação Mestrado - Tiago Barros Santos Malta.pdf: 8207831 bytes, checksum: a5e3ee5357f988c47072c9481326377e (MD5) Previous issue date: 2016-02-25 / CAPES, CNPQ, FACEPE / O câncer de mama feminino é o segundo tipo de câncer mais frequente no mundo, com aumento de incidência de 22% a cada ano. Estudos nas últimas décadas revelaram que a transformação maligna está associada a uma variedade de células com padrões de glicosilação alterados, como por exemplo a sialilação. Os ácidos siálicos tem sido relacionados à iniciação e progressão do câncer, tendo assim implicações potenciais na prevenção, diagnóstico e tratamento da doença. Este trabalho avaliou a expressão fenotípica de ST3Gal-I, através da imunohistoquímica, e o perfil de ácido siálico, através da histoquímica com lectinas usando Maackia Amurensis agglutinin II (MAA), em tecidos mamários diagnosticados com fibroadenoma (n=59), carcinoma ductal in situ (CDIS, n=40), carcinoma ductal invasivo (CDI, n=50) e carcinoma lobular (CL, n=42). Todos os tipos de lesões de mama apresentaram alta imunopositividade à ST3Gal-I, sendo observada uma expressão em 93,2% dos casos de Fibroadenoma, 92,5% de CDIS, 96% dos casos CDI e 85,2% de CL. As células apresentaram um padrão de marcação citoplasmático e perinuclear com relação à ST3Gal-I. Diferentes distribuições de resíduos de ácido siálico ⍺2,3-ligados, com um padrão de marcação predominantemente citoplasmático e membranar, foram observados nas lesões de mama estudadas. Os casos de fibroadenoma apresentaram o menor percentual de sialilação entre as lesões analisadas (47,5%), enquanto os de CDI o maior pecentual (98%). Embora este estudo não tenha mostrado diferença significativa na expressão de ST3Gal-I entre as lesões de mama, alterações representativas na presença de ácidos siálicos entre fibroadenoma e lesões malignas (p<0,0001), e também entre CDIS e CDI (p = 0.037) foram notadas. Não foram encontradas correlações significativas entre as expressões de ST3Gal-I e MAA, os marcadores de rotina e as características clinico-histopatológicas dos pacientes. Os resultados indicam uma distribuição particular de ácidos siálicos ⍺2,3-ligados nas células mamárias entre as lesões estudadas sugerindo seu envolvimento na progressão/manutenção do câncer de mama. / The female breast cancer is the second most common type of cancer in the world, with increased incidence of 22% every year. Recent studies in recent decades have revealed that the malignant transformation is associated with a variety of cells with altered glycosylation patterns such as sialylation. Sialic acids have been demonstrated to participate in cancer initiation and progression, thus has potential implications for cancer prevention, diagnosis and treatment. This study evaluated the phenotype expression of ST3Gal-I using immunohistochemistry and the sialic acid residues profile by histochemistry with Maackia amurensis agglutinin II (MAA) in mammary tissues diagnosed as fibroadenoma (n=59), ductal carcinoma in situ (DCIS, n=40), invasive ductal carcinoma (IDC, n=50) and lobular carcinoma (LC, n=42). All types of breast lesions showed high ST3Gal-I immunopositivity, its expression was observed in 93.2% cases of Fibroadenoma, 92.5% of DCIS, 96% of IDC and 85.2% cases of LC. The cells ST3Gal-I staining pattern was mainly cytoplasmatic and perinuclear. The MAA staining in breast lesions showed a diffuse cytoplasmatic and membrane pattern with different distribution of ⍺2,3-linked sialic acids among the lesions studied, fibroadenoma cases showed the lowest percentage among the analyzed lesions (47.5%) while IDC showed the highest (98%). Although this study did not show a significant difference in expression of ST3Gal-I among all lesions, representative alterations in sialic acid content between fibroadenoma and malignant lesions (p<0.0001), and also between CDIS and CDI (p=0.037) were observed. No significant correlations were found between the expressions of ST3Gal-I and MAA, routine markers and clinical-histopathological characteristics of the patients. Results indicate different distribution of ⍺2,3-linked sialic acids on the cells of the studied lesions which seems to be involved in breast cancer progression and/or maintenance.

Sialyltransferases, CMP-sialic acid synthetases and CMP-sialic acid transport proteins play a crucial role in the construction of cell surface glycoconjugates. These proteins also have a pivotal role to play in a number of diseases, including cancer. The sialyltransferase enzymes are responsible for transfering sialic acids from the donor substrate (CMP-sialic acid) to growing cell surface glycoconjugate chains within the Golgi apparatus. The CMP-sialic acid synthetase enzymes are responsible for the synthesis of the CMP-sialic acid, the donor substrate of the sialyltransferases in the nucleus, while the CMP-sialic acid transport proteins are responsible for transporting CMP-sialic acid from the Cytosol to the Golgi apparatus. When these proteins function in an abnormal way, hypersialylation results, leading to an increased level of sialylation on the cell surface. This increased level of sialylation aids in the detachment of primary tumour cells due to an increase in the level of overall negative charge, causing repulsion between the cancer cells. Therefore, the sialyltransferase enzymes, CMP-sialic acid synthetases and CMP-sialic acid transport proteins are intimately involved in the metastatic cascade associated with cancer. Chapter 1 provides a general introduction of cancer metastasis, discussing the roles of three target proteins (CMP-sialic acid synthetases, CMP-sialic acid transport proteins and sialyltransferases), as well as discussing their substrate specificities, with an emphasis on their involvements in cancer metastasis. The Chapter concludes with an overview of the types of compounds intended to be utilised as probes or inhibitors of these proteins. Chapter 2 describes the general approach towards the synthesis of CMP-Neu5Ac mimetics with a sulfur linkage in the presence of a phosphate group in the general structure 38. The precursor phosphoramidite derivative 45 was prepared and isolated in a good yield using Py.TFA. Unfortunately, the target compound 38 could not be prepared. Chapter 3 describes an alternative strategy wherein S-linked sialylnucleoside mimetics, of the general structure 39, with a sulfur linkage, but no phosphate group, between the sialylmimetic and the ribose moiety in the base is targeted. A series of these S-linked sialylnucleoside mimetics were successfully prepared. Cytidine, uridine, adenosine and 5-fluorouridine nucleosides were used to create a library of different nucleosides and with structural variability also present in the sialylmimetic portion. This small 'library' of 15 compounds was designed to shed light on the interaction of these compounds with the binding sites of the sialyltranferase, CMP-sialic acid synthetase and/or CM-sialic acid transport protein. Approaches towards the synthesis of O-linked sialylnucleoside mimetics of the general structure 40 are described in Chapter 4. Several methodologies are reported, as well as protecting group manipulations, for successful preparation of these sialylnucleoside mimetics. Cytidine and uridine were employed as the nucleosides, thus allowing a direct comparison between the O- and S-linked sialylnucleoside mimetics in biological evaluation. It appears from these synthetic investigations that gaining access into the O-linked series is not as straightforward as for the S-linked series, with alternative protecting group strategies required for the different nucleosides. The biological evaluation of some of the compounds reported in Chapters 3 and 4 is detailed in Chapter 5. The sialylnucleoside mimetics were evaluated, by 1H NMR spectroscopy, for their ability to inhibit CMP-KDN synthetase. In addition, an initial 1H NMR spectroscopic-based assay was investigated for inhibition studies of α(2,6)sialyltranferase in the absence of potential inhibitors. The final chapter (Chapter 6) brings together full experimental details in support of the compounds described in the preceding Chapters.

Dissertation (Ph.D.)--University of South Florida, 2008. / Title from PDF of title page. Document formatted into pages; contains 140 pages. Includes vita. Includes bibliographical references.

Submitted by Caroline Falcao (caroline.rfalcao@ufpe.br) on 2017-04-04T19:50:14Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) 2012-Dissertação-SteffanyFerreira.pdf: 961397 bytes, checksum: 6fda526203d2fb7ff12a32289be35454 (MD5) / Made available in DSpace on 2017-04-04T19:50:14Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) 2012-Dissertação-SteffanyFerreira.pdf: 961397 bytes, checksum: 6fda526203d2fb7ff12a32289be35454 (MD5) Previous issue date: 2012 / O câncer de pele ou tumor cutâneo não-melanoma (TCNM), é a neoplasia maligna de maior incidência no Brasil. Os cânceres, em geral, apresentam padrão de glicosilação aberrante na estrutura de oligossacarídeos de superfície celular (fenótipo), determinada pela ação conjunta de glicosiltransferases (genótipo). O presente estudo objetiva avaliar o perfil glicofenotípico de glicoconjugados de superfície celular de TCNM e correlacionar com o padrão de expressão de sialiltransferases. Cortes (4µm) de biópsias de Carcinoma Basocelular (CBC), Carcinoma Espinocelular (CEC), Ceratose Actínica (CA) e Ceratoacantoma (KA) foram ensaiados com as lectinas SNA, Sambucus nigra agglutinin, (específica para ácidos siálicos ligados na posição α-2,6) e MAA, Maakia amurensis agglutinin (específica para ácidos siálicos na posição α-2,3). A inibição da ligação lectina-carboidrato foi realizada com o ácido siálico (300mM). As amostras também foram avaliadas com anticorpos monoclonais anti-ST3Gal I (específico para β-galactosideo α2,3 sialiltransferase) e antiST6Gal I (específico para β-galactosideo α2,6 sialiltransferase). Semelhanças no padrão de sialilação entre CEC, CA e KA foram encontradas na histoquímica com lectinas e, principalmente na imunohistoquímica. O CBC não apresentou marcação na maioria dos casos para ambas as técnicas, indicando que este tipo tumoral apresenta um padrão molecular próprio podendo estar associado ao seu comportamento biológico (baixo potencial invasivo e metastático). Os resultados indicam a importância da sialilação no desenvolvimento e manutenção de tumores cutâneos, sendo a histoquímica com lectinas e a imunohistoquímica ferramentas úteis na determinação deste tipo de glicosilação na superfície de células tumorais. / Skin cancer or Non-melanoma Cutaneous Tumors (NMCT) is the most incidente malign neoplasia in Brazil. Cancers, in general, present a aberrant glycosilation pattern in oligossacharides of cell surfasse glycoconjugates (phenotype) which it is determined by tha action of glycosiltransferases (genotype). The present study aims to evaluate the glycophenotype of glycoconjugates in the cell surface of NMCT and to correlates to the expression of sialyltransferases. Biopsies slices (4µm) of basal cell carcinoma (BCC), squamous cell carcinoma (SCC), actinic keratosis (AK) and Keratoacanthoma (KA) were assayed with SNA, Sambucus nigra agglutinin, (specific to α2,6- linked sialyc acid) and MAA, Maakia amurensis agglutinin (specific to α2,3- linked sialic acid). Samples were also evaluated with monoclonal antibodies anti-ST3Gal I (specific to Sialyltransferase β-galactoside α2,3-linked) and antiST6Gal I (specific to Sialyltransferase β-galactoside α2,6-linked). Sialylation patterns were similar to SCC, AK and KA in both lectin histochemistry and immunohistochemistry but mainly in the later one. BCC was not positive in most of the cases for both methods which indicates that this tumour has a particular molecular pattern that can be associated to its biological behavior (low invasive and metastatic potential). Results indicate the importance of silaylation in the development and maintenance of cutaneous tumors being the lectin histochemistry associated to the immunohistochemistry a useful auxiliary tool for the determination of this type of glycosilation in skin tumors.

Dissertation (Ph.D.)--University of South Florida, 2008. / Includes vita. Also available online. Includes bibliographical references (leaves 121-140).

Este trabajo demuestra que los genes que codifican para los enzimas beta-galactosido alfa-2,3-sialiltransferasa 3 (ST3Gal III), y en menor medida beta-galactosido alfa-2,3-sialiltransferasa 4 (ST3Gal IV), están directamente implicados en etapas clave de la progresión tumoral como la adhesión, la migración y la formación de metástasis en las líneas de adenocarcinoma pancreático humano Capan-1 y MDAPanc-28. También, que las Especies Reactivas del Oxígeno (ROS) generadas durante los procesos de proliferación y diferenciación celular o debido a estímulos oxidantes externos, desempeñan un importante papel en el control de la síntesis de ST3Gal III y SLex, y por lo tanto en la regulación del fenotipo metastático. Además, junto al papel pro-adhesivo de la E-Selectina, este trabajo ha descrito efectos prometastáticos adicionales para esta molécula como inductora de la migración y de la secreción de VEGF a través de un mecanismo E-Selectina-SLex dependiente. / This work shows that genes that codifying for the enzymes beta-galactoside alpha-2,3-sialyltransferase 3 (ST3Gal III), and in a lower extent beta-galactoside alpha-2,3-sialyltransferase 4 (ST3Gal IV) , are directly implicated in key steps of tumour progression such as adhesion, migration and metastasis formation in the pancreatic adenocarcinoma cell lines Capan-1 and MDAPanc-28. Moreover, Reactive Oxygen Species (ROS) generated in these cell lines during cell proliferation-differentiation processes or by external oxidant stimuli, play a role in the control of ST3Gal III and SLex levels and in the acquisition of a more aggressive phenotype. And, together with the pro-adhesive role of E-Selectin for circulating cells, this work uncovers sE-Selectin dependent migration and VEGF secretion through a SLex depending mechanism, supporting additional pro-metastatic effects for sE-Selectin-SLex interaction.

Thesis (Ph.D.)--University of Alabama at Birmingham, 2008. / Title from first page of PDF file (viewed on June 29, 2009). Includes bibliographical references.