published_or_final_version / abstract / Medicine / Master / Master of Philosophy

Thesis (Ph. D.)--University of Alabama at Birmingham, 2006. / Title from first page of PDF file (viewed Oct. 31, 2007). Includes bibliographical references.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 4 uppsatser.

Diss. (sammanfattning) Stockholm : Karol. inst., 2005. / Härtill 6 uppsatser.

Diss. (sammanfattning) Stockholm : Karol. inst., 2005. / Härtill 5 uppsatser.

Diss. (sammanfattning) Stockholm : Karol inst., 2004. / Härtill 4 uppsatser.

Thesis (Ph.D. in Biophysics & Genetics) -- University of Colorado Denver, 2007. / Typescript. Includes bibliographical references (leaves 118-124). Free to UCD affiliates. Online version available via ProQuest Digital Dissertations;

Thesis (Ph. D.)--Case Western Reserve University, 2005. / [School of Medicine] Department of Molecular Biology and Microbiology. Includes bibliographical references. Available online via OhioLINK's ETD Center.

Thesis (doctoral)--Rijksuniversiteit te Utrecht.

nÃo hà / IntroduÃÃo: MutaÃÃes nos genes GNAS e AIP estÃo presentes em 35% e 3%, respectivamente, dos somatotrofinomas esporÃdicos. Recentemente, observa-se importÃncia biolÃgica crescente dos microRNAs (miRNAs) na tumorigÃnese hipofisÃria. Entretanto, mecanismos moleculares envolvidos na patogÃnese de 60% desses tumores permanecem nÃo elucidados. Objetivos: Identificar a prevalÃncia de mutaÃÃes nos genes GNAS e AIP em um grupo de somatotrofinomas esporÃdicos. Comparar parÃmetros clÃnicos e bioquÃmicos ao diagnÃstico como idade, tamanho tumoral e agressividade (escore Hardy), hormÃnio do crescimento (GH), prolactina (PRL) e Fator de Crescimento Insulin-Like I (IGF-1) e resposta as terapÃuticas entre os grupos com (gsp+) e sem (gsp-) mutaÃÃo no GNAS. Analisar a expressÃo dos genes GNAS e BTG2 e miRNAs entre somatotrofinomas e hipÃfises normais (HN) e a associaÃÃo entre a expressÃo com agressividade, a resposta à cirurgia e a todas as terapÃuticas adjuvantes disponÃveis. Material e MÃtodos: 26 pacientes com diagnÃstico de acromegalia. Tamanho tumoral foi avaliado por RNM/CT e o grau de invasibilidade pelo escore de Hardy (I a IV). GH basal ≤2.5μg/L ou nadir de GH apÃs o GTT≤1μg/L e IGF-1 normal para idade e sexo foram utilizados como critÃrio de cura apÃs cirurgia transesfenoidal (CTE). Como controle com o anÃlogo da somatostatina (AS), adotamos a normalizaÃÃo dos nÃveis de IGF-1 para idade e sexo. As amostras tumorais (n=26) foram obtidas durante a CTE, realizado histopatolÃgico e armazenadas a -70 ÂC, para estudos moleculares. HN (07) foram obtidas durante autÃpsias. RNA e DNA total foram extraÃdos pelo TRIzolÂ. Os cÃdons 201 e 227 do gene GNAS e o AIP completo foram sequenciados. ExpressÃo relativa dos genes GNAS e BTG2 e dos miRNAs let-7a, miR-16a, miR-21, miR-141, miR-143, miR-15a, miR-145, miR-23a, miR-23b e miR-24-2 foi avaliada por qPCR (sondas TaqMan), pelo mÃtodo 2-ΔΔCt. Resultados: A frequÃncia de mutaÃÃes no GNAS foi de 35% e no AIP 3,8%. NÃo houve diferenÃa entre as mÃdias de idade (39,0Â11,5 vs 43,6Â9,0 anos; p=0,32), nas concentraÃÃes plasmÃticas basais de GH (62,4Â128,1 vs 39,9Â48,3Âg/L; p=0,39), IGF-1 (435,5Â230,8 vs 556,9 238,3 %ULNR; p=0,32), PRL (25,7Â29,8 vs 30,9Â32,8 ng/L; p=0,69) e agressividade tumoral entre os gsp+ e gsp-(p=1,00). Ao analisar o uso do AS como terapÃutica adjuvante à CTE, observamos que 04/05 (80%) dos indivÃduos com somatotrofinoma gsp+ obtiveram controle da doenÃa, enquanto que no grupo gsp- 02/06 (33%) obtiveram controle (p=0,08). Quando associamos ao AS, os agonistas dopaminÃrgicos e/ou radioterapia externa, observamos que 05/05 (100%) dos pacientes gsp+ tiveram critÃrio de controle da doenÃa, contra (04/09) 44% no grupo gsp- (p=0,09). NÃo houve diferenÃa na expressÃo de GNAS entre os somatotrofinomas e as HN (1,07Â0,55 vs 0,98Â0,28; p=0,97), e entre os gsp+ e gsp- (1,04Â0,59 vs 1,10Â0,55; p=0,97, respectivamente). Os tumores Hardy I / II apresentaram maior expressÃo do GNAS do que os tumores classificados como III / IV (p=0,02). NÃo houve associaÃÃo entre a expressÃo do GNAS e o controle da doenÃa com cirurgia isolada ou com o uso de todas as terapÃuticas adjuvantes. Observamos hipoexpressÃo do BTG2 e dos miR-16a e miR-141 em somatotrofinomas quando foram comparados com as HN (p=0,002, fold=-6,63; p=0,01, fold=-10,00; p=0,0003, fold=-50,00, respectivamente) sem diferenÃas entre os gsp+ e gsp-. Houve hiperexpressÃo do miR-21 (p=0,02;fold=10,18) em somatotrofinomas (20,16Â18,48) quando comparado com as HN (2,52 Â3,56), sem diferenÃa entre os gsp + e gsp-. NÃo houve diferenÃa na expressÃo entre os grupos gsp+ e gsp- para os miRNAs let-7a, miR-21, miR-143, miR-15a, miR-23a e miR-24-2. Entretanto, miR-145 e miR-23b foram mais hipoexpressos no grupo gsp+ quando comparados ao gsp- (p=0,03, fold=-4,83 e p=0,02, fold=-2,77, respectivamente). NÃo houve associaÃÃo entre a expressÃo do BTG2 e o painel de miRNAs com agressividade e com o controle da doenÃa. ConclusÃo: Na presente sÃrie de somatotrofinomas, assumidos como esporÃdicos, a frequÃncia de mutaÃÃes nos genes GNAS (35%) e AIP (3,8%) foram semelhantes aos relatados na literatura. NÃo houve diferenÃas nas caracterÃsticas clÃnicas e bioquÃmicas, agressividade, resposta Ãs terapÃuticas, e na expressÃo diferencial do GNAS entre os pacientes com tumores gsp+ e gsp-. HipoexpressÃo de BTG2 (gene supressor tumoral relacionado Ãs vias de sinalizaÃÃo do p53 e do Rb), baixa expressÃo de miRNAs (supressores tumorais) e alta expressÃo de oncomirs em somatotrofinomas sugerem um papel desses na tumorigÃnese somatotrÃfica. / Introduction: Mutations in GNAS and AIP genes are present in 35% and 3%, respectively, of the sporadic somatotropinomas. Recently, increased biological importance of microRNAs (miRNAs) has been observed in pituitary tumorigenesis. However, the molecular mechanisms involved in the pathogenesis of 60% of these tumors remain to be elucidated. Objectives: To identify the prevalence of mutations in GNAS and AIP genes in a series of sporadic somatotropinomas. Compare clinical, bioquimical parametrer at diagnosis as age, tumor size and theirs aggressiveness, pre-operative growth hormone (GH), prolactin (PRL) and insulin-like growth factor-I (IGF-1) levels and treatment responsiveness between somatotropinomas with (gsp+) and without (gsp-) GNAS mutation.To analyze the expression of GNAS and BTG2 genes and a panel of miRNAs between somatotrofinomas and normal pituitaries (NP) and the association between the expression of these genes and miRNAs with aggressiveness, as well as disease control with surgery or control with all adjuvant therapeutic approaches. Material and Methods: 26 patients with acromegaly. GH basal ≤2.5μg/L or nadir after OGTT ≤1μg/L and normal IGF-I matched for age and sex were used as diagnosis and for cure criteria after transsphenoidal surgery (TS). As control after somatostatin analogues (SA), we adopted the normalization of IGF-I matched for age and sex. Tumor size was evaluated by MRI/CT and the degree of invasiveness by Hardy score (I to IV).Tumor samples (26) were obtained during TS, processed for histopathology and stored at -70ÂC for molecular studies. NP (07) were obtained during autopsy. Total DNA and RNA were extracted by TRIzolÂ. Codons 201 and 227 of the GNAS gene and the whole AIP gene were sequenced. Relative expression of BTG2 and GNAS genes and miRNAs let-7a, miR-16a, miR-21, miR-141, miR-143, miR-15a, miR-145, miR-23a, miR-23b, and miR-24-2 was measured by qPCR (TaqMan probes) using 2-ΔΔCt method. Results: Frequencies of GNAS and AIP mutations were 35% and 3.8%, respectively. There was no difference between the mean age (39.0  11.5 vs 43.6  9.0 years, p=0.32), basal GH (62.4Â128.1 vs 39.9  48.3 μg/L; p=0.39), IGF-I (435.5  230.8 vs. 556.9  238.3; p=0.32) and PRL (25.7  29.8 vs. 30.9  32.8 ng/L, p=0.69) in plasma concentration, and tumor aggressiveness (p=1.00) between (gsp+) and (gsp-) groups. We observed that 80% (04/05) of gsp+ whereas 33% (02/06) of the gsp- achieved control (p=0.07) after SA therapy adjuvant to TS. When SA, dopamine agonists and/or external radiotherapy were associated 100% (05/05) of gsp+ group and 44% (04/09) of gsp- group (p=0.08) showed disease control.There was no difference in GNAS expression between somatotropinomas and NP (1.07  0.55 vs 0.98  0.28, p=0.97) as well as between somatotropinomasgsp+ and gsp- (1.04  0.59 vs 1.10  0.55, p=0.97, respectively). Hardy I/II tumors showed higher GNAS expression than Hardy III/IV (p=0.02), but there was no association between GNAS expression and disease control with surgery alone or associated with other adjuvant therapies. We observed hypoexpression of BTG2 and miR-16a and miR-141 in somatotropinomas compared with NP (-6.6 fold, p=0.002; -10.0 fold, p=0.01; and -50.0 fold, p=0.0003, respectively) with no difference between gsp+ and gsp- somatotropinomas. There was miR-21 overexpression in somatotropinomas compared with NP (20.2  18.5 vs 2.5  3.6; 10.2 fold, p=0.02), with no difference between gsp+ and gsp- somatotropinomas. However, miR-145 and miR-23b were more hipoexpressed in gsp+ compared to gsp- (-4.8fold, p=0.03 and-2.7 fold, p=0.02). There was no association between the expression of BTG2 and a panel of miRNAs with aggressiveness or disease control. Conclusion: In this series of assumed sporadic somatotopinomas, the frequencies of mutations in GNAS (35%) and AIP (3.8%) were similar to the literature. There were no differences in clinical and biochemical characteristics, aggressiveness, response to therapy, and GNAS expression in patients with gsp+ and gsp- somatotropinomas. Hypoexpression of BTG2, a tumor suppressor gene related to p53 and Rb signaling pathways, low expression of tumor suppressor miRNAs and high expression of oncomirs in somatotropinomas suggest a role in the somatotrophic tumorigenesis.