Thesis (M.Sc.)--Memorial University of Newfoundland, Faculty of Medicine, 2000. / Typescript. Bibliography: leaves 92-100.

Thesis (Ph.D.)--Memorial University of Newfoundland, Faculty of Medicine, 2001. / Typescript. Bibliography: leaves 188-241.

Made available in DSpace on 2014-12-17T15:32:23Z (GMT). No. of bitstreams: 1 AndreiaFC_DISSERT.pdf: 3035785 bytes, checksum: a9769f0a294924c551036d1a098b7d2a (MD5) Previous issue date: 2014-02-12 / A displasia epitelial (DE) oral ? uma desordem potencialmente maligna (DPM), cujo diagn?stico e grada??o histol?gica se baseiam nas suas altera??es arquiteturais e citol?gicas. Para avaliar o risco de transforma??o maligna dessas les?es de forma mais precisa ? fundamental entender e localizar altera??es gen?ticas e epigen?ticas nas c?lulas displ?sicas, as quais podem ajudar a compreender melhor a progress?o para a malignidade. Dessa forma, o presente estudo objetivou avaliar a imunoexpress?o de EGFR e PTEN nas DEs orais e relacionar esse aspecto com as caracter?sticas cl?nicas e grada??o histol?gica pelo sistema bin?rio (baixo e alto risco de transforma??o maligna). Para tanto, foram selecionados 20 casos de DE de alto risco e 20 de baixo risco para serem submetidos ? an?lise imunoistoqu?mica para os biomarcadores supracitados. A imunomarca??o de cada caso foi avaliada semiquantitativamente atrav?s de escores e quanto ? localiza??o nos estratos epiteliais. A an?lise estat?stica foi realizada atrav?s dos testes de Mann-Whitney, Qui-quadrado de Pearson, Exato de Fisher e de correla??o de Spearman com n?vel de signific?ncia estabelecido em 5%. Os resultados mostraram que 57,5% dos pacientes eram do g?nero feminino, a m?dia de idade foi de 57,5 anos, 42,5% foram diagnosticados clinicamente como leucoplasia e a maioria dos casos foi proveniente de les?es localizadas na l?ngua (32,5%). De forma geral, g?nero e idade n?o exerceram influ?ncia na imunoexpress?o do EGFR e PTEN. A express?o do EGFR foi observada em 100% dos casos, nos quais houve predom?nio do escore 3 (75%) e imunoreatividade em todas as camadas epiteliais (55%), independente da grada??o histol?gica (p = 0,453 e p = 0,204, respectivamente). O PTEN revelou positividade de marca??o em 87,5% dos casos, nos quais observou-se predom?nio do escore 0 (55%) e imunoreatividade limitada ? camada basal (40%), por?m sem diferen?as significativas entre os grupos histol?gicos (p = 0,904 e p = 0,915, respectivamente). Por fim, quando analisados, em conjunto, os 40 casos de DEs, foi observada uma fraca correla??o positiva, estatisticamente significativa, entre os padr?es de imunoexpress?o do EGFR e do PTEN (r = 0,317; p = 0,046). Com base nesses resultados, altera??es no padr?o de express?o do EGFR e PTEN sugerem que essas prote?nas participam de processos moleculares relacionados com a carcinog?nese em mucosa oral

Nutrient and growth factor receptors internalize through dathrin coated pits. The signal sequences which mediate the association between receptors and the coated pit reside in receptor cytoplasmic tail domains. These signal sequences have been extensively investigated in nutrient receptors, and a minimal functional sequence has been identified consisting of a tyrosine residue in an exposed b turn. Protein-protein contacts between internalization signal sequences and components of the coated pit machinery have been proposed to mediate rapid internalization. In vitro evidence suggests the AP-2 adaptor may be that protein component. The signal sequences of growth factor receptors are less well understood. However, a growth factor- and temperature- dependent binding between the epideimal growth factor receptor and the AP-2 adaptor has been observed. We identified Hsc70 as a cytosolic ligand for the cytoplasmic tail of the transferrin receptor. The binding was mapped to the internalization signal sequence of the receptor tail. Mutations within the signal sequence which inhibit internalization result in alteration of signal sequence secondary structure and reduction in stimulation of the Hsc70 ATPase. Co-immunoprecipitation analysis showed a population of transferrin receptors which are bound to Hsc70, suggesting an association in vivo. We also showed binding of Hsc70 to the epidermal growth factor receptor by co-immunoprecipitation analysis. This binding was increased by treatment with EGF. The binding was transient, and occured prior to the binding of the receptor to AP-2 adaptors. Other agents which induce EGF receptor clustering and internalization also stimulate the transient increase in Hsc70 binding and the later AP-2 binding, suggesting a role in early endocytosis. These data support the hypothesis that Hsc70 is associated with the receptors for transferrin and epidermal growth factor in vitro and in vivo. We propose a role for the 70 kD heat shock protein in the assembly/disassembly of protein complexes involved in receptor signalling and/or internalization.

The altered protein expression and activity of receptor tyrosine kinases (TK) are implicated in the progression of various types of cancers. One such dysfunction is the overexpression of the epidermal growth factor receptor (EGFR) that correlates with aggressive tumor progression and poor prognosis. On the other hand, c-Src non-receptor tyrosine kinase is overexpressed and activated in a large number of human malignancies and has been strongly linked to progression to distant metastases. c-Src-induced phosphorylation of EGFR is required for EGF-mediated mitogenesis, tumorigenesis and tumour invasiveness. Thus we surmised that molecules termed "combi-molecules" designed to block both EGFR and c-Src should not only possess significant growth inhibitory potency but also strong anti-invasive properties. In this thesis, we utilized molecular modeling to design molecules containing two moieties: one that straddles the structure of the known Src inhibitor PP2 and the other that mimics the backbone of Iressa, a potent EGFR inhibitor. Of all the molecules synthesized, only SB163 containing the longest spacer between the two moieties was capable of inducing a dose dependent inhibition of both Src and EGFR. More importantly, SB163 blocked cell motility in the wound healing assay and showed significantly greater anti-invasive activity than a PP2+Iressa combination. The observation that SB163 was a less potent EGFR or Src inhibitor than Iressa and PP2 suggests that its superior potency when compared with the PP2+ Iressa combination may be at least partially attributed to mechanisms other than EGFR or Src blockade. This was also corroborated by the fact that SB163, despite its significant bulkiness (>700) could induce dose dependent inhibition of other kinase such PDFGR and Abl. The results in toto suggest that conferring multiple kinase targeting properties to single molecules can lead to highly anti-proliferative and anti-invasive agents. Traditionally, multi-kinase targeted molecules were discovered serendipitously through multi-kinase testing. Here we initiated a more rational approach to the design of single multi-targeted molecules. Cancer being a complex disease driven by tumours characterized by multiple disordered signaling pathways, this approach may well represent a novel avenue in the therapy of refractory malignancies.

Dissertation (Ph.D.)--University of South Florida, 2005. / Includes vita. Includes bibliographical references. Also available online in PDF format.

Diss. (sammanfattning) Umeå : Umeå universitet, 2008. / Härtill 4 uppsatser.

Orientador: Luis Otavio Zanatta Sarian / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-28T02:45:23Z (GMT). No. of bitstreams: 1 Baptista_MauricioZuccolotto_D.pdf: 4237946 bytes, checksum: b677fee05d5d510e473e6b4c4c856a53 (MD5) Previous issue date: 2015 / Resumo: Introdução: A via de sinalização intracelular PTEN, mTOR, PI3K, IGF-1R e EGFR exerce papel prognóstico importante no câncer de mama. Embora muitos estudos tenham analisado a correlação entre as alterações gênicas de PTEN, mTOR, PI3K, IGF-1R e EGFR e o mau prognóstico em câncer de mama, há uma lacuna na literatura com relação ao valor prognóstico dessas proteínas na adjuvância do câncer de mama. Apesar de existirem fatores prognósticos e preditivos conhecidos, como os receptores hormonais e o HER-2, no campo imunológico, o câncer de mama é um dos tumores menos imunogênicos. As proteínas PD-L1 e PD-L2 fazem parte de uma importante resposta imune antitumoral. Em câncer de mama, o valor prognóstico de PD-L1 e PD-L2 ainda não está definido. Objetivos: Investigar a expressão das proteínas PTEN, mTOR, PI3K, IGF-1R e EGFR e das proteínas PD-L1 e PD-L2, e a correlação com as características clínicas e patológicas do câncer de mama, sobrevida livre de doença e sobrevida global. Métodos: Foi realizada uma coorte que avaliou 192 casos de câncer de mama, estadios I, II e III, tratadas entre 1994 e 2014 no Hospital da Mulher (CAISM) da UNICAMP. Os dados clínicos e de sobrevida foram retirados de prontuários. Blocos de parafina foram utilizados para construção do microarranjo de tecidos (TMA). No TMA foi utilizada a técnica de imunohistoquímica (IHQ) para estudo da expressão dessas proteínas. A terapia adjuvante foi administrada de acordo com o protocolo de tratamento institucional. Resultados: A expressão de PTEN foi encontrada em 40.6% (77/190); mTOR em 47.4% (90/190); PI3K em 29.8% (57/191); IGF-1R em 35.8% (68/190) e EGFR em 25.7% (49/191). Nas células do câncer de mama, a expressão de PTEN ocorreu no citoplasma e na região nuclear; a expressão de mTOR foi constatada de modo intenso na região nuclear e também no citoplasma. A expressão de PI3K foi mais intensa na membrana celular e menos intensa no citoplasma. A expressão de IGF-1R foi detectada na membrana celular das células tumorais. A expressão de todas essas proteínas foi significativamente associada à presença de linfonodos positivos. A idade mais jovem ao diagnóstico foi associada à expressão de PTEN e PI3K. A presença de tumores maiores foi associada à expressão de PTEN. Os receptores de progesterona negativos foram associados à expressão de PI3K. Receptores de estrógeno negativos e recorrência à distância foram ambos associados à expressão de EGFR. As expressões de PTEN, PI3K e EGFR foram fortemente associadas a características clínicas e patológicas de pior prognóstico. A expressão de PI3K foi significativamente associada à pior sobrevida livre de progressão (p=0.04) e pior sobrevida global (p=0.04). A expressão de EGFR foi também significativamente associada à pior sobrevida livre de progressão (p=0.03) e pior sobrevida global (p=0.04). A expressão de PTEN não foi associada à sobrevida. A expressão de PD-L1 foi identificada em 56.7% (107/189) e a de PD-L2 em 50.8% (97/192). Enquanto a expressão de PD-L1 foi detectada na membrana celular e no citoplasma das células do câncer de mama, a expressão de PD-L2 ocorreu no citoplasma e na região nuclear. Idade mais jovem ao diagnóstico, linfonodos positivos, receptor de estrógeno negativo e recorrência à distância foram associados tanto à expressão de PD-L1 quanto à de PD-L2. A presença de tumores maiores e de alto grau histológico foi associada à expressão de PD-L1. A expressão de PD-L1 foi significativamente associada à melhor sobrevida global (p=0.04). Conclusão: A expressão de PTEN, PI3K e EGFR pode representar um tipo de câncer de mama mais agressivo. A expressão de PI3K e EGFR pode ser considerada um marcador de mau prognóstico em câncer de mama. A expressão de PD-L1, apesar de ser associada a características clínicas e patológicas de pior evolução, pode ser considerada um marcador de bom prognóstico em câncer de mama / Abstract: Introduction: The PTEN, mTOR, PI3K, IGF-1R and EGFR signaling pathway plays an important role in prognosis of breast cancer. Although many studies have analyzed the correlation between PTEN, mTOR, PI3K, IGF-1R, and EGFR genes alterations with poor prognosis in breast cancer, there is still a gap in the literature concerning the prognostic value of these proteins in the adjuvant setting. Despite there were some well-known predictive and prognostic factors, such as hormone receptors and HER-2, in the immune field, breast cancer is one of the less immunogenic tumors. PD-L1 and PD-L2 constitute an important antitumor immune response. In breast cancer, the prognostic value of PD-L1 and PD-L2 is still to be defined. Objectives: This study investigates PTEN, mTOR, PI3K, IGF-1R and EGFR proteins expression and PD-L1 and PD-L2 proteins expression, and their correlation with clinicopathological features, disease-free survival and overall survival. Methods: In order to assess these proteins expression, we conducted an immunohistochemistry study using a tissue microarray encompassing 192 breast cancer cases, stage I, II and III, treated between 1994 and 2014 at the Women¿s Hospital (CAISM) from UNICAMP. All clinical and outcome data were retrieved from medical charts. Adjuvant therapy was administered according to the institution¿s treatment protocol. Results: PTEN expression was found in 40.6% (77/190); mTOR expression in 47.4% (90/190); PI3K expression in 29.8% (57/191); IGF-1R expression in 35.8% (68/190); and EGFR expression in 25.7% (49/191). In breast cancer cells PTEN expression showed cytoplasmic and nuclear immunoreactivity, and mTOR expression revealed strong nuclear and cytoplasmic staining. Tumors harboring PI3K expression presented strong immunoreactivity at cell membrane and weak cytoplasmic staining. IGF-1R expression was detected in breast cancer cell membrane. All proteins expression was significantly associated with lymph node positivity. Younger age at diagnosis was related to PTEN and PI3K expression. The presence of larger tumors was associated with PTEN expression. Negative progesterone receptor was correlated to PI3K expression. Estrogen receptor negativity and recurrence at distant sites were associated with EGFR expression. The expression of PTEN, PI3K, and EGFR were strongly associated with clinical and pathological features of poor prognosis. In our cohort, PI3K expression was associated with significantly worse disease-free survival (p=0.04) and overall survival (p=0.04), and EGFR expression was also significantly associated with worse disease-free survival (p=0.03) and overall survival (p=0.04). PD-L1 expression was present in 56.7% (107/189), and PD-L2 expression was identified in 50.8% (97/192). In breast cancer cells PD-L1 expression revealed strong immunoreactivity at cell membrane and cytoplasmic staining, and PD-L2 expression showed cytoplasmic and nuclear immunoreactivity. Younger age at diagnosis, lymph node positivity, estrogen negative receptor, and recurrence at distant sites were all associated with both PD-L1 and PD-L2 expression. The presence of larger tumors and higher histological grade were both associated with PD-L1 expression. PD-L1 expression was significantly associated with better overall survival (p=0.04) in breast cancer patients. Conclusion: The expression of PTEN, PI3K, and EGFR can represent a more aggressive type of breast cancer. PI3K and EGFR expressions emerge as poor prognostic markers in breast cancer. Despite its association with poor clinical and pathological features, PD-L1 expression seems to be a good prognostic marker in breast cancer / Doutorado / Oncologia Ginecológica e Mamária / Doutor em Ciências da Saúde

No description available.

<p>Receptor epidermalnog faktora rasta (EGFR) pripada porodici receptora protein-tirozin kinaze čija je aktivacija povezana sa proliferacijom malignih, invazijom, inhibicijom apoptoze, tumorskom angiogenezom i metastatskim &scaron;irenjem stoga ima važnu ulogu u karcinogenezi i tumorskoj progresiji. Aktivirane mutacije se odvijaju oko katalitičkog tirozin kinaza domena. Biopsijski, citolo&scaron;ki i hirur&scaron;ki uzorci se koriste u detekciji EGFR mutacija u momentu postavljanja dijagnoze adenokarcinoma ili karcinoma sa komponentom adenokarcinoma, najpouzdanije lančanom reakcijom polimeraze. Činjenica da primena ciljane molekularne terapije tirozin kinaza inhibitorima kod obolelih sa EGFR mutiranim adenokarcinomom pluća pobolj&scaron;ava prognozu bolesti, postoji rezistencija kod pojedinih tipova EGFR mutacija i povezanost histopatolo&scaron;kim i imunohistohemijskim karakteristikama tumora, da je bronholo&scaron;ki uzorak često jedini uzorak u kome je potrebno odrediti i molekularni profil tumora osnovni cilj ove disertacije bio je da se odredi učestalost i tip EGFR mutacija i povezanost sa karakteristikama adenokarcinoma. Da bi se taj cilj realizovao postavljeni su sekundarni ciljevi odnosno da se: izvr&scaron;i histopatolo&scaron;ka reklasifikacija adenokarcinoma pluća na osnovu kriterijuma koje je postavila internacionalna asocijacija za proučavanje carcinoma pluća, američko torakalno dru&scaron;tvo i evropsko respiratorno dru&scaron;tvo; odredi ekspresija TTF-1 u adenokarcinomu pluća i povezanost sa EGFR mutacionim statusom; odredi učestalost, tip i povezanost EGFR mutacija sa predominantnim tipom adenokarcinoma i utvrdi da li bronhoskopska biopsija može da bude reprezentativni uzorak za određivanje EGFR mutacionog statusa. Histopatolo&scaron;ka građa adenokarcinoma pluća u hirur&scaron;kim uzorcima je heterogenija u odnosu na biopsijske uzorke i ta razlika je statistički značajna (p&lt;0,001). Acinarno predominantni tip je najzastupljeniji u hirur&scaron;kim i biopsijskim uzorcima bez statistički značajne razlike u raspodeli predominantnih tipova u njima (p=0,65883). Predominantni tip u primarnom tumoru određuje predominantni tip u limfogenim metastazama. EGFR mutacije tipa insercija na egzonu 21 i L858R mutacija na egzonu 20 su detektovane kod tri od 60 (5%) bolesnika u pet od 120 uzoraka (tri hirur&scaron;ka i dva biopsijska uzorka), če&scaron;će kod žena, starijih od 60 godina, pu&scaron;ača i u solidno predominantnom tipu. Ne postoji statistički značajna razlika u koncentraciji izolovane DNK između EGFR mutiranih i wt EGFR adenokarcinoma u biopsijskim (p=0,132) i hirur&scaron;kim uzorcima (p=0,641). Procenat invalidnih rezultata prilikom određivanja EGFR mutacionog statusa u je veći u biopsijskim uzorcima u odnosu na hirur&scaron;ke uzorke. Postoji statistički značajna razlika izmeĐu broja TTF-1 pozitivnih i TTF-1 negativnih adenokarcinoma (p&lt;0,001), ali ne i u raspodeli ovih bolesnika prema polovima (p=0,1231), prosečnoj starosti, pu&scaron;ačkim navikama (p=0,6488) i prosečnoj veličini tumora (p=0,21). Postoji pozitivna korelacija između TTF-1 pozitivne ekspresije i EGFR mutacionog statusa stoga TTF-1 pozitivna ekspresija može da bude prediktor pozitivnog EGFR mutacionog statusa. Bronhoskopska biopsija je reprezentativni uzorak za određivanje EGFR mutacionog statusa zato &scaron;to: većina dijagnostičkih biopsijskih uzoraka ima vi&scaron;e od 100 očuvanih tumorskih ćelija, nema razlike u raspodeli predominantnih tipova u odnosu na hirur&scaron;ke uzorke, EGFR mutacije se detektuju u uzorcima sa manje od 100 tumorskih ćelija i manje od 20% volumenske gustine tumorskog tkiva, razlika između koncentracije izolovane DNK u EGFR mutiranim i wt EGFR adenokarcinomima u biopsijskim i hirur&scaron;kim uzorcima nije statistički značajna (p=0,132 i p=0,641).</p> / <p>Epidermal growth factor receptor (EGFR) belongs to the family of protein-tyrosin kinase family, whose activation is associated with the proliferation of malignant cells, invasion, inhibition of apoptosis, tumor angiogenesis and metastatic spread and thus plays an important role in carcinogenesis and tumor progression. Activated mutations take place around the catalytic tyrosine kinase domain. Biopsy, cytological and surgical specimens are used for the detection of EGFR mutations at the time of diagnosis of adenocarcinoma or carcinoma with an adenocarcinoma component, most reliably using a polymerase chain reaction. The fact that the application of molecular tyrosin kinase inhibitor therapy to patients with EGFR mutated lung adenocarcinoma improves the prognosis of the disease, there is resistance in certain types of EGFR mutations and connection with histopathological and immunohistochemical characteristics of tumor, that the bronchoscopic specimen is often the only specimen in which it is necessary to determine the molecular profile of the tumor, the primary objective of this thesis is to determine the frequency and type of EGFR mutations and their connection with the characteristics of adenocarcinoma. In order to realize this objective, the following secondary objectives have been set: to execute histopathological reclassification of lung adenocarcinoma based on the criteria set by the International Association for the Study of Lung Cancer, the American Thoracic Society and European Respiratory Society; determine the expression of TTF-1 in lung adenocarcinoma and connection with EGFR mutation status; determine the frequency, type and connection of EGFR mutations with predominant type of adenocarcinoma and confirm whether bronchoscopic biopsy may be a representative specimen for the determination of EGFR mutation status. Histopathological material of lung adenocarcinoma in surgical specimens is more heterogeneous in relation to biopsy specimens and such difference is statistically significant (p&lt;0,001). Acinar predominant type is the most common in surgical and biopsy specimens with no statistically significant differences in the distribution of predominant type among them (p=0,65883). The predominant type in the primary tumor determines the predominant type in lymphatic metastases. EGFR mutations in the type of insertions on exon 21 and L858R mutations on exon 20 have been detected in three out of 60 (5%) of patients in five out of 120 specimens (three surgical and two biopsy samples), more often in women older than 60, smokers and in a solid predominant type. There are no statistically significant differences in the concentration of isolated DNA between EGFR mutated and wt EGFR adenocarcinoma in biopsy (p=0,132) and surgical specimens (p=0,641). The percentage of invalid results in determining the EGFR mutation status is higher in biopsy specimens compared to the surgical specimens. There is a statistically significant difference between the number of TTF-1 positive and TTF-1 negative adenocarcinoma (p&lt;0,001), but not in the distribution of these patients according to gender (p=0,1231), average age, smoking habits (p=0,6488) and average tumor size (p-0,21). There is a positive correlation between TTF-1 positive expression and EGFR mutation status and therefore TTF-1 positive expression can be a predictor of positive EGFR mutation status. Bronchoscopic biopsy is a representative sample for the determination of EGFR mutation status because: most diagnostic biopsy specimens have more than 100 preserved tumor cells, there is no difference in the distribution of predominant types in relation to surgical specimens, EGFR mutations are detected in samples with less than 100 tumor cells and less than 20% of volume density of tumor tissue, the difference between the concentration of isolated DNA in EGFR mutated and wt EGFR adenocarcinoma in biopsy and surgical specimens is not statistically significant (p=0,132 and p=0,641).</p>