O καρκίνος του μαστού αποτελεί την δεύτερη πιο συχνή μορφή καρκίνου που απαντάται και στα δύο φύλα ενώ είναι συνιστά την πιο συχνά εμφανιζόμενη μορφή καρκίνου στις γυναίκες καθώς και την πρώτη αιτία θανάτου στο γυναικείο φύλο. Η γενετική ανάλυση του κληρονομικού καρκίνου έχει ενταχθεί τα τελευταία χρόνια και αποτελεί σημαντικό μέρος της κλινικής πράξης, καθώς όλο και περισσότεροι ασθενείς ελέγχονται για την ύπαρξη γαμετικών μεταλλάξεων στα υψηλής διεισδυτικότητας γονίδια BRCA1 και BRCA2. Προκειμένου να μελετήσουμε την συνεισφορά των γονιδίων BRCA1 και BRCA2 στην εμφάνιση καρκίνου μαστού στον ελληνικό πληθυσμό εξετάστηκαν 200 άτομα που παρουσίαζαν σημαντικό ατομικό ή/και οικογενειακό ιστορικό καρκίνου του μαστού ή/και των ωοθηκών. Έτσι, στην περίπτωση του γονίδιου BRCA1 βρέθηκαν 13 διαφορετικές παθογόνοι μεταλλάξεις σε 29 από τις ασθενείς (14.5%) ενώ για το γονίδιο BRCA2 αναλύθηκαν συνολικά 156 ασθενείς (155 γυναίκες και 1 άνδρας) και μεταλλάξεις ανιχνεύθηκαν σε 8 γυναίκες (5.2%) και 1 άνδρα (100%). Παράλληλα έγινε προσπάθεια χαρακτηρισμού του φάσματος μεταλλάξεων του μετρίου διεισδυτικότητας γονιδίου RAD51C σε ελληνικές οικογένειες. Η ανάλυση αυτή πραγματοποιήθηκε σε 87 ασθενείς όμως δεν ανιχνεύθηκε κάποια παθογόνος μετάλλαξη σε κάποιον ασθενή. Επιπρόσθετα, πραγματοποιήθηκε η ανίχνευση μεγάλων γονιδιακών αναδιατάξεων στα γονίδια BRCA1 και BRCA2 με την βοήθεια τριών τεχνικών ανίχνευσης: την τεχνική της ανίχνευσης γονιδιακών αναδιατάξεων με χρήση διαγνωστικών εκκινητών για τις πιο συχνές γονιδιακές αναδιατάξεις του γονιδίου BRCA1 στον ελληνικό πληθυσμό (δύο απαλειφές του εξωνίου 20, μία απαλειφή των εξονίων 23 και 24 και τέλος μια απαλειφή του εξονίου 24), την τεχνική της πολλαπλής και ποσοτικής αλυσιδωτής αντίδρασης της πολυμεράσης μικρών φθοριζόντων τμημάτων (QMPSF) για το γονίδιο BRCA1 και την τεχνική της πολλαπλής ενίσχυσης τμημάτων DNA μέσω ιχνηθετών (MLPA®) τόσο για το γονίδιο BRCA1 όσο και για το γονίδιο BRCA2. Η τεχνική της ανίχνευσης με την χρήση διαγνωστικών εκκινητών εφαρμόστηκε σε 200 ασθενείς με καρκίνο μαστού ή/και ωοθηκών και ανιχνεύθηκαν 15 ασθενείς που έφεραν κάποια από τις συχνότερες γονιδιακές αναδιατάξεις του γονιδίου BRCA1 (7.5%). Αντίθετα, με την τεχνική του QMPSF και του MLPA δεν ανιχνεύθηκε κάποια γονιδιακή αναδιάταξη στα BRCA1/2 στους συνολικά 24 ασθενείς που εξετάστηκαν. Επιπλέον, στα πλαίσια της παρούσας διατριβής πραγματοποιήθηκε η ανάλυση 403 γυναικών με τριπλά αρνητικό καρκίνο του μαστού ανεξαρτήτως οικογενειακού ιστορικού και ηλικίας εμφάνισης της νόσου. Κατά την ανάλυση αυτή ελέγχθηκαν συγκεκριμένα εξόνια καθώς και γονιδιακές αναδιατάξεις του BRCA1. Μεταλλάξεις ανιχνεύθηκαν σε 65 από τις ασθενείς αυτές (16%). Στα πλαίσια της παρούσας διατριβής πραγματοποιήθηκε ανάλυση απλοτύπου προκειμένου να αποδειχθεί ότι η απαλειφή των εξονίων 23 και 24 αποτελεί ελληνική ιδρυτική μετάλλαξη. Για το σκοπό αυτό αναλύθηκε ο απλότυπος σε 21 οικογένειες που έφεραν την συγκεκριμένη γονιδιακή αναδιάταξη με την βοήθεια της τεχνικής των πολυμορφικών δεικτών επαναλήψεων μικροδορυφορικού DNA. Η ανάλυση απλοτύπου κατέδειξε ότι 8 από 10 γενετικούς τόπους είναι κοινοί μεταξύ των ατόμων που φέρουν την απαλοιφή επιβεβαιώνοντας έτσι την υπόθεση ότι η απαλειφή αυτή αποτελεί ιδρυτική μετάλλαξη του ελληνικού πληθυσμού. Ακολούθως, μελετήθηκε η συνεισφορά και άλλων γονιδίων στην προδιάθεση για καρκίνο μαστού και ωοθηκών και πραγματοποιήθηκε ανάλυση 21 γονιδίων (BRCA1, BRCA2, CHEK2, PALB2, BRIP1, TP53, PTEN, STK11, CDH1, ATM, BARD1, MLH1, MRE11, MSH2, MSH6, MUTYH, NBN, PMS1, PMS2, RAD50, RAD51C) με την χρήση τεχνολογίας αλληλούχισης επόμενης γενεάς. Για το σκοπό αυτό αναλύθηκαν 42 ασθενείς με σοβαρό ιστορικό καρκίνο μαστού ή/και ωοθηκών, οι οποίες είχαν βρεθεί αρνητικές για μεταλλάξεις στα γονίδια BRCA1 & 2. Κατά την ανάλυση αυτή βρέθηκαν μεταλλάξεις σε 11 ασθενείς (26.2%). Ακόμα, πραγματοποιήθηκε εξομική αλληλούχιση σε μια ασθενή, η οποία παρουσίαζε βαρύτατο ατομικό και οικογενειακό ιστορικό και είχε βρεθεί αρνητική για μεταλλάξεις σε 21 γονίδια (BRCA1, BRCA2, CHEK2, PALB2, BRIP1, TP53, PTEN, STK11, CDH1, ATM, BARD1, MLH1, MRE11, MSH2, MSH6, MUTYH, NBN, PMS1, PMS2, RAD50, RAD51C). Η αλληλούχιση αυτή πραγματοποιήθηκε από την εταιρεία BGI. Συνολικά ανιχνεύθηκαν 39.762 πολυμορφισμοί ενός νουκλεοτιδίου (SNPs) και 1,646 μικρές ενθέσεις/απαλειφές (InDels) σε ολόκληρο το γονιδίωμα της ασθενούς. Μετά από ανάλυση των αποτελεσμάτων αυτών καταλήξαμε σε 12 παραλλαγές που αποτελούν ενδεχομένως πιθανά παθογόνα αλληλόμορφα. / Breast cancer is the second most common cancer in both sexes while it is the most frequent cancer as well as the first cause of death in women. The genetic testing of hereditary cancer is part of the everyday clinic as the number of patients that is checked for germline mutation in BRCA1 and BRCA2 is continuously increasing. In order to study the contribution of the high penetrance genes BRCA1 and BRCA2 in breast cancer in Greece 200 patients with severe history of breast and/or ovarian cancer were screened. In BRCA1 13 different pathogenic mutations were found in 29 patients (14.5%) while in BRCA2 156 patients were screened (155 female and 1 male) and pathogenic mutations were identified in 8 female (5.2%) and 1 male patients (100%). At the same time, we tried to characterize the mutation spectrum of the medium penetrance gene RAD51C in Greek families. For this purpose, we screened 87 patients but no mutations was found. In addition, the identification of large genomic rearrangements in both genes took place, using three methods: diagnostic primers were used in order to identify the 4 most common BRCA1 genomic rearrangements in Greece (two deletions in exon 20, one deletion of exons 23 and 24 and one in exon 24), the QMPSF method for the BRCA1 genomic rearrangements (Quantitative Multiplex PCR of Short Fluorescent fragments) and the MLPA method for both BRCA1 and BRCA2 genomic rearrangements (Multiplex Ligation-dependent Probe Amplification). With the first technique 200 patients with breast and/or ovarian cancer were analyzed and 15 of them were found to carry one of the most frequent BRCA1 genomic rearrangements. In the contrary, no large genomic rearrangements in neither gene was identified among the 24 patients who were screened using the two latter methods (QMPSF and MLPA). Also, we have screened 403 triple negative female patients who were diagnosed with triple-negative invasive breast cancer, independently of their age or family history, for germline BRCA1 mutations in exons where a mutation have previously been found in Greek population including the genomic rearrangements. Mutation were identified in 65 patients (16%). In this study we used haplotype analysis in order to demonstrate that the BRCA1 deletion of exons 23 and 24 constitutes a Greek founder mutation. For this purpose, we performed haplotype analysis in 21 Greek families who carry this specific genomic rearrangement using the DNA Microsatellite Repeats analysis. The haplotype analysis showed that 8 out of 10 genetic regions are common among people that carry this deletion confirming that the deletion is a Greek founder mutation. We also studied the contribution of other genes in breast/ovarian cancer predisposition by studying 21 genes (BRCA1, BRCA2, CHEK2, PALB2, BRIP1, TP53, PTEN, STK11, CDH1, ATM, BARD1, MLH1, MRE11, MSH2, MSH6, MUTYH, NBN, PMS1, PMS2, RAD50, RAD51C) using next generation sequencing techniques. We examined 42 patients with severe breast and/or ovarian cancer history that were found negative for BRCA1/2 mutations. In this analysis pathogenic mutation were found in 11 patients (26.2%). At last, exome sequencing was performed in a female patient who had severe family history and was found negative for germline mutations in 21 genes (BRCA1, BRCA2, CHEK2, PALB2, BRIP1, TP53, PTEN, STK11, CDH1, ATM, BARD1, MLH1, MRE11, MSH2, MSH6, MUTYH, NBN, PMS1, PMS2, RAD50, RAD51C). The sequencing was carried out by Beijing Genomics Institute (BGI). In total, were identified 39.762 singe nucleotide polymorphisms (SNPs) and 1.646 insertions - deletions (InDels) in the patient's exome. After the result analysis we concluded in 12 variants that could constitute possible pathogenic alleles.

No description available.

Thèse numérisée par la Division de la gestion de documents et des archives de l'Université de Montréal

Diss. (sammanfattning) Stockholm : Karol. inst., 2006. / Härtill 4 uppsatser.

Thesis (Ph. D.)--University of Washington, 2007. / Vita. Includes bibliographical references (leaves 116-125).

Source: Masters Abstracts International, Volume: 46-04, page: 2093. Advisers: Xianglin L. Du; Karen H. Lu. Includes bibliographical references.

Dissertation (Ph.D.) --University of Texas Graduate School of Biomedical Sciences at San Antonio, 2008. / Vita. Includes bibliographical references.

The ovarian surface epithelium (OSE) is a monolayer of cells surrounding the ovary that is ruptured during ovulation. After ovulation the wound is repaired, however this process, and the mechanisms to maintain OSE homeostasis after the wound is repaired are poorly understood. We have shown the mouse OSE (mOSE) contains a stem cell population that is expanded by Transforming Growth Factor Beta 1 (TGFB1), a factor present in follicular fluid. These data suggest that components in the follicular fluid such as TGFB1 may promote wound repair and OSE homeostasis through maintenance of the OSE stem cell population. Additionally, TGFB1 may promote wound repair through induction of an epithelial-to-mesenchymal transition (EMT) and activation of pro-survival pathways, as seen in other tissues. To elucidate the mechanism for TGFB1-mediated ovulatory wound repair, mOSE cells were treated with TGFB1, which induced an EMT seen with increased Snai1 expression and cell migration. Snai1 overexpression also increased cell migration and sphere formation (a stem cell characteristic). RNA sequencing results suggest this is at least in part through elevated collagen deposition in SNAI1 overexpressing cells. A TGFB signalling targets array identified Cox2 induction following TGFB1 treatment. Constitutive Cox2 expression did not promote an EMT, but enhanced sphere formation and cell survival. Finally, TGFB1 treatment decreased Brca1 expression, which when deleted from mOSE cells also increased sphere formation. RNA sequencing results suggest that Brca1 deletion promotes stemness through activation of the stem cell genes Ly6a and Lgr5. RNA sequencing was also used to compare mOSE cells cultured as monolayers and as spheroids, with and without TGFB1. These results validate our findings that TGFB1 promotes an EMT partially through Snail induction and the upregulation of Cox2. mOSE cells cultured as spheroids acquire a mesenchymal transcriptional profile that is further enhanced with TGFB1 treatment. These data suggest that TGFB1 may promote ovulatory wound repair and maintain OSE homeostasis through the induction of an EMT, maintenance of the stem cell population and activation of a pro-survival pathway. Interestingly, mOSE spheroids also decrease Brca1 expression and upregulate cancer associated genes such as Pax8 and Greb1. The induction of survival pathways, while simultaneously increasing stemness and repressing Brca1 could render cells more susceptible to transformation. This work provides novel insights as to why ovulation is the primary non-hereditary risk factor for ovarian cancer.

O câncer de mama é uma das neoplasias malignas mais comuns que afetam mulheres de todo o mundo. No Brasil, o Estado do Rio Grande do Sul tem índices de incidência e mortalidade por câncer de mama que situam-se entre os maiores do país. Aproximadamente 5-10% dos diagnósticos são causados por mutações germinativas em genes de predisposição entre os quais estão BRCA1 e BRCA2, associados à Síndrome de Câncer de mama e Ovário Hereditários (Hereditary Breast and Ovarian Cancer Syndrome ou HBOC, OMIM #114480).A identificação dos casos hereditários de câncer de mama é importante porque indivíduos afetados apresentam risco cumulativo vital muito superior ao da população para o desenvolvimento de câncer, porque familiares de um afetado podem estar igualmente em risco porque há medidas de rastreamento intensivo e intervenções preventivas que podem diminuir significativamente o risco de câncer em portadores de mutação. O diagnóstico molecular da síndrome HBOC é laborioso e caro devido à heterogeneidade molecular da doença. Famílias que apresentam características indicativas de uma síndrome de predisposição ao câncer de mama e ovário hereditários, mas que são negativas para mutações pontuais em BRCA1/2 vêm sendo testadas para grandes rearranjos visto que essas anormalidades têm sido consideradas como respondendo por, no mínimo, 10% do todos os casos HBOC com mutação identificável, incluindo grandes deleções ou duplicações. Um estudo recente de Portugal, demonstrou que um rearranjo fundador no exon 3 de BRCA2 ocorre em por 8% das famílias HBOC do Norte do país. Os objetivos deste trabalho incluíram a verificação da freqüência e caracterização de rearranjos gênicos nos genes BRCA1 e BRCA2, incluindo a mutação fundadora c.156_157insAlu no exon 3 de BRCA2 em famílias brasileiras dealto risco para a síndrome HBOC. Em um grupo de 145 indivíduos em risco nãorelacionados rastreados para a mutação fundadorac.156_157insAlu no exon 3 de BRCA2 foram encontrados 3 portadores da mutação (prevalência de 2%). Em um grupo de 145 indivíduos de risco não-relacionados rastreados para rearranjos gênicos em BRCA1 e BRCA2 pela técnica de MLPA (multiplex ligation-dependent probe amplification) foram identificados 4 portadores de mutação germinativa, sendo a mutação em dois deles um rearranjo gênico no gene BRCA1 (1,4%) envolvendo sequencias Alu. Rearranjos gênicos em BRCA1 e BRCA2 são responsáveis por uma parcela das mutações em famílias HBOC Brasileiras. O presente estudo, envolvendo uma série grande de famílias com o fenótipo da síndrome HBOC, não identificou novos rearranjos fundadores, no entanto, demonstrou a presença de rearranjos tanto em BRCA1 quanto em BRCA2, reiterando a importância da busca ativa por estas alterações, que dificilmente são identificadas por técnicas convencionais de sequenciamento gênico. A técnica de MLPA associada a um protocolo específico para detecção da mutação fundadora Portuguesa c.156_157insAlu podem ser utilizadas como estratégia inicial de rastreamento de mutações em famílias Brasileiras com a síndrome. Os resultados apresentados aqui, no entanto, indicam que mutações serão identificadas em menos de 10% dos casos utilizando esta estratégia. / Breast cancer is one of the most common malignancies affecting women worldwide. In Brazil, the State of Rio Grande do Sul has incidence rates and mortality from breast cancer are among the largest in the country. Approximately 5-10% of the cases are caused by germline mutations in predisposing genes including BRCA1 and BRCA2 are associated with the syndrome of breast and ovarian cancer Hereditary (Hereditary Breast and Ovarian Cancer Syndrome or HBOC, OMIM # 114480). The identification of inherited cases of breast cancer is important because affected individuals have cumulative risk life much higher than the population for developing cancer because of an affected family may also be at risk because there are measures of intensive screening and preventive interventions that can significantly decrease the risk of cancer in mutation carriers. The molecular diagnosis of HBOC syndrome is laborious and expensive due to the molecular heterogeneity of the disease. Families that have characteristics indicative of a cancer predisposition syndrome of hereditary breast and ovarian cancers, but are negative for mutations in BRCA1/2 have been tested for large rearrangements because these abnormalities have been identified as accounting for at least 10 % of all cases HBOC identifiable mutation, including large deletions or duplications. A recent study from Portugal, the founder showed that a rearrangement in exon 3 of BRCA2 occurs in 8% of HBOC families of the north. The objectives of this work included the verification of the frequency and characterization of gene rearrangements in BRCA1 and BRCA2 genes, including c.156_157insAlu founder mutation in exon 3 of BRCA2 mutations in Brazilian families at high risk for HBOC syndrome. In a group of 145 individuals at risk unrelated traced to c.156_157insAlu founder mutation in exon 3 of 3 found BRCA2 mutation carriers (prevalence 2%). In a group of 145 individuals at risk unrelated screened for gene rearrangements in BRCA1 and BRCA2 by the technique of MLPA (multiplex ligationdependent probe amplification) identified four carriers of germline mutation, and two of the mutation in a gene rearrangement in the gene BRCA1 (1.4%) involving Alu sequences. Gene rearrangements in BRCA1 and BRCA2 account for a portion of HBOC mutations in Brazilian families. This study, involving a large series of families with HBOC syndrome phenotype, no new rearrangements identified founders, however, showed the presence of rearrangements in both BRCA1 and BRCA2, reiterating the importance of active search for these changes, which hardly are identified by conventional techniques of gene sequencing. The technique of MLPA protocol associated with a specific mutation detection founder Portuguese c.156_157insAlu strategy can be used as initial screening for mutations in families with Brazilian syndrome. The results presented here, however, indicate mutations that will be identified in less than 10% of the cases using this strategy.

Det finns flera olika typer av en bröstcancergen, två av dessa cancergener är BRCA 1 och 2. Dessa cancergener är ärftliga och risken för att utveckla bröst eller livmodercancer blir upp till 80 % när man är bärare redan vid 30 års ålder. Syftet är att beskriva unga kvinnors upplevelser av att leva med genmutation BRCA 1 och/eller BRCA 2. Studien är en litteraturstudie. Dataanalysen mynnade ut i fyra teman känsla av hotad existens, behov av stöd för att hantera information, avsaknad av samhörighet och etiskt dilemma. Resultatet lyfter fram de upplevelser och fruktan som är central för unga kvinnor när de får besked om att vara bärare av BRCA 1 eller 2 och de beslut de måste ta som kan påverka livet och framtiden. Studiens resultat är användbart för vårdpersonal och närstående som möter dessa unga kvinnor och kan ge en större förståelse för känslor och upplevelser som uppstår som bärare på en cancergen.