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The pathogenesis of megaloblastic hemopathies (MH) is centered on the deficiency of vitamin B12 and folic acid with interruption of erythrocyte maturation. This study researched the participation of p53 and p21 in the pathophysiology of the disease. A retrospective study enrolled 95 patients with histopathologic diagnosis by biopsy or bone marrow clot (BMB/BMC), with clinical review and immunohistochemical study in tissue microarray (TMA) for p53 and p21, detailing their marking location. All patients had BMC and only 11 had BMB. The CMO was a differential of this study and it allowed an expanded sample. In the TMA, 63.7% (58/91) of the samples were immunopositive for p53; and 35.2% (31/88) were immunopositive for p21. Nuclear staining, divergent from the literature, was observed in 17.3% (10/58) among those p53+ and in 38.7% (12/31) among those p21+. The pattern of immunostaining showed non-significant differences (P=0.474) regarding morphologic and clinical aspects. The positivity for both may indicate an effective balance between apoptosis and anti-apoptotic action. Excessive inhibition of apoptosis would contribute to high global cellularity, but without functional maturation effectiveness. In conclusion, there is p21 and/or p53 immunoexpression in most cases of this study and there is no clear association between immunoexpression pattern and patient outcome. Unlike the literature, we also found a percentage of nuclear immunostaining, but the finding was not statistically significant. Combination of p21 and p53 results created different possibilities of pathologic interpretation for MH, reinforcing the importance of studies similar to this one.

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The aim of the present study was to determine the clinical significance of p53 and p21ras p21wafl, p27kip1 and p16ink4a expression in cases of early gastric cancer. A total of 81 patients who had undergone gastrectomy with D2 lymphadenectomy between 1971 and 2004 were retrospectively investigated. The immunohistochemical expression of p21ras, p53, p21waf1/cip1, p27kip1 and p16ink4a in the tissues was evaluated. In normal, metaplastic and tumoral mucosa, p53 was positive in 53, 87.3, and 87.1% of the cases, respectively. In the same tissues, p21ras was positivE in 85.3, 86 and 96.8%, respectively. Positivity FOR p16ink4a was DETECTED IN 46.3, 91.1 and 86% OF THE CASES, respectively, WHEREAS p27kip1 WAS positiVE IN 60, 94.7 and 95.3%, and p21wafl/cip1 WAS positivE IN 32.4, 72.7 and 71.4% OF THE CASES, respectively. All THE tumors WERE positive for p53. Tumors with lymph node invasion presented WITH OVERexpression (+4) of p53 in 47% of the cases VS. 17% OF patients who DID not HAVE lymph node involvement. THEREFORE, higher expression of p53, p21ras and p21wafl/cip1 IN the tumor exhibited a statistically significant association with lymph node involvement.

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Background: In colorectal cancer, BRAF and KRAS mutation are mutually exclusive, but both are independent prognostic factors for the disease. Aim: To determine the frequency of BRAF V600E mutation in colorectal cancer. Material and Methods: A KRAS mutation study was carried out in 100 tissue samples of primary and metastatic adenocarcinomas of colon and rectum from patients aged 61.1 ± 62 years (56 women). Negative KRAS mutation cases underwent study of BRAF V600E mutation by restriction fragment length polymorphism (RFLP) and direct sequencing. Results: Primary tumors were located in the colon and rectum in 88 and six cases respectively. Five were liver metastases and in one case, the sample location was undetermined. Forty two samples were KRAS positive (mutated). In 12 of the 58 KRAS negative (wild type) samples, the V600E mutation in codon 15 of the BRAF gene was demonstrated. No differences in the frequency and distribution of mutations, stratified by gender, age, primary tumor versus metastasis, or tumor location were observed. Conclusions: Twelve percent of KRAS negative colorectal cancer samples showed BRAF gene mutation. Considering that 42% of samples have a KRAS mutation, 54% of patients should not respond to therapies with monoclonal antibodies directed against epidermic growth factor (EGFR) pathway.

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A incidência do melanoma cutâneo em pacientes adultos jovens tem aumentado consideravelmente nos últimos anos. Há, contudo, carência de conhecimentos clinicopatológicos e moleculares sobre os melanomas que ocorrem nessa faixa etária. O presente estudo teve por objetivo avaliar 132 casos de melanoma cutâneo primário em pacientes com idade entre 18 e 30 anos, com ênfase no estudo das características clínicas, histopatológicas e avaliação molecular das mutações nos genes BRAF, NRAS e KIT. Em relação aos achados clínicos e histopatológicos, houve predomínio de indivíduos do sexo feminino (61,4%), sendo o tronco o sítio anatômico mais comumente envolvido (44,3%) e o melanoma extensivo superficial o tipo histológico predominante (79,5%). A mutação V600E no gene BRAF (BRAFV600E) foi analisada em 93 casos, utilizando-se a técnica de RT-PCR. Essa mutação foi identificada em 38,7% (36/93) e, estatisticamente, associada à fase vertical de crescimento (p = 0,01), infiltrado inflamatório discreto (p = 0,02) e presença de mitose intradérmica (p = 0,004). Houve, ainda, forte indício de associação com a presença de ulceração (p = 0,05). Todas essas variáveis apresentaram associação com pior prognóstico do melanoma cutâneo. Observou-se predomínio da mutação BRAFV600E em regiões anatômicas relacionadas à exposição solar intermitente. Nenhum caso de melanoma com fenômeno de regressão apresentou mutação BRAFV600E (p < 0,05). Não houve associação significativa entre BRAFV600E e sexo, tipo histológico, nível de Clark, índice de Breslow, elastose solar, invasão angiolinfática e perineural, satelitose, nevo melanocítico coexistente e sobrevida. A pesquisa de mutações NRAS, pela técnica de RT-PCR, detectou frequência de 3,95% (3/76). As três mutações encontradas foram do tipo 61K e ocorreram em pacientes do sexo masculino e em região de cabeça e pescoço. As mutações BRAFV600E e NRAS, quando presentes, eram mutuamente exclusivas. A frequência de mutações KIT, analisadas por...

The incidence of cutaneous melanoma in young adults has dramatically increased in recent years. However, there is scarce data about the clinicopathological and molecular characteristics on the melanomas occurring at this age group. The present study aimed to evaluate 132 patients aged between 18 and 30 years with primary cutaneous melanoma with emphasis on the study of clinical, histopathological characteristics and molecular evaluation of mutations in BRAF, NRAS and KIT genes. Regarding the clinical and histopathological findings, the following results were found: female predominance (61.4%), trunk was the most commonly anatomical site involved (44.3%) and superficial spreading melanoma, was the most common histological type (79.5 %). The V600E mutation in BRAF (BRAFV600E) gene was analyzed in 93 cases, using RT-PCR. It was present in 38.7% (36/93) and statistically related to the vertical growth phase (p = 0.01), mild inflammatory infiltration (p = 0.02) and the presence of intradermal mitosis (p = 0.004). There was, also, strongly evidence of an association with the presence of ulceration (p = 0.05). Worse prognosis was associated with these variables. There was a predominance of BRAFV600E mutation in anatomical regions related to intermittent sun exposure. No cases of melanoma with BRAFV600E mutation showed regression phenomenon (p < 0.05). There was no significant association between BRAFV600E and gender, histological type, Clark level, Breslow thickness, solar elastosis, angiolymphatic and perineural invasion, sattelitosis, coexisting melanocytic nevus and survival. The presence of a mutation in NRAS, by RT-PCR was seen in 3.95% (3/76) of the cases. All these three mutations were of type 61K, occurred in male patients and the head and neck region. BRAFV600E and NRAS mutations, when present, were mutually exclusive. The frequency of KIT mutations, analyzed by sequencing, was 11.1% (3/27). The three mutations identified in this gene were located in...

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BACKGROUND: KRAS gene mutations play an important role in the carcinogenesis of colorectal tumors. However, studies that have assessed the association between KRAS gene mutation status and disease characteristics report conflicting results. To assess KRAS gene status (mutated or wild-type) and its association with the clinical, epidemiological, and histopathological features of metastatic colorectal adenocarcinoma as well its association with clinical outcomes. METHODS: Cross-sectional descriptive study in which clinical and histopathological data were collected from the medical records of 65 patients diagnosed with metastatic colorectal adenocarcinoma at the Clinical Oncology Service of the Teaching Hospital of the School of Medicine of Ribeirao Preto, University of Sao Paulo (Hospital das Clinicas da Faculdade de Medicina de Ribeirao Preto, Universidade de Sao Paulo -HCFMRP-USP) between 2005 and 2012 and analyzed based on their KRAS gene status. RESULTS: KRAS gene mutations were found in 49.2% of the tumors, and G/A (25.5%) and Gly12Asp (34.37%) were the most frequent mutations. Among the investigated clinical features (gender, ECOG (Eastern Cooperative Oncology Group), histology, degree of cell differentiation, lymph node ratio, primary tumor site, staging, presence of synchronous metastasis, lung metastasis, and liver metastasis), the association between age less than 65 years with KRAS mutation was statistically significant (P = 0.046). KRAS mutation status did not exhibit a significant correlation with the overall survival of the patients (P = 0.078); however, the cases with KRAS mutation exhibited shorter survival. In the multivariate analysis, synchronous metastasis (P = 0.03) and liver metastasis (P = 0.008) behaved as independent factors of poor prognosis relative to the overall survival of the patients. CONCLUSION: The KRAS mutation status did not exhibit prognostic value in the investigated sample. Among the older patients (> 65 years old), wild-type KRAS was more frequently observed compared to mutated KRAS.

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Introdução: Cerca de 20% a 40% da população apresenta nódulos tireoidianos assintomáticos e, aproximadamente 30% destes nódulos apresentam malignidade ao exame anatomopatológico pós-cirúrgico. Tentativas de se estabelecer associações biológicas e diferenças entre os distúrbios benignos e malignos da tireóide mostram resultados conflitantes. Estudos prévios utilizando cDNA microarrays identificaram genes com expressão diferencial em diferentes patologias tireoidianas, incluindo IGFBP5; proteases como a catepsina B e inibidores de proteinases como a a1 antitripsina; componentes envolvidos nos processos de endocitose e transporte intracelular como a clatrina e; reguladores do ciclo celular como p27 e p21. Objetivo: analisar, caracterizar e comparar o perfil de expressão protéico de seis genes diferencialmente expressos em tireóide normal, bócio, adenoma folicular e carcinomas diferenciados e, determinar a importância clínica dos fenótipos identificados. Métodos: 172 amostras de tecido tireoidiano incluindo 18 normais, 18 bócios, 50 adenomas foliculares, 50 carcinomas papilíferos e 36 carcinomas foliculares foram coletadas em duplicata para análise e comparação dos padrões de expressão de seis proteínas (p27, p21, IGFBP5, clatrina, a1-antitripsina e catepsina B) utilizando técnicas de imunoistoquímica em tissue microarrays. Os casos foram avaliados de acordo com a intensidade da coloração (O a 4) e porcentagem de células positivas (O a 4). Resultados: Foi observada expressão diferencial para alfa 1 antitripsina, clatrina e IGFBP5, p21 e p27 quando os tecidos foram comparados (p< 0,0001 ). Tecidos neoplásicos exibem expressão aumentada de IGFBP5 em relação aos não neoplásicos. Nenhuma diferença significativa foi observada em relação à expressão de catepsina B. Nas células foliculares normais e de bócio, a imunorreatividade para p27 e p21 foi predominantemente nuclear em comparação aos carcinomas. Este mesmo padrão foi observado em adenoma quando comparados aos carcinomas foliculares (p< 0,008). Carcinomas papilíferos exibiram expressão citoplasmática aumentada de p27 e p21. Conclusão: Vários e novos genes detectados por técnicas de análise genômica em larga escala podem estar envolvidos na patogênese molecular e progressão tumoral. A transcrição destes genes pode resultar na superexpressão de proteínas como IGFBP5, clatrina e alfal antitripsina, enquanto modificações pós-transcricionais resultam em alteração da expressão de proteínas nucleares envolvidas no controle do ciclo celular e, que contribuem para o processo de malignização da tireóide. Dados obtidos em TMA são consistentes com os resultados obtidos por cDNA microarray mostrando expressão protéica diferencial para p27, p21, IGFBP5, clatrina e al-antitrpsina em tecido tireoidiano normal, hiperplásico e neoplásico; o que poderia ser de valor diagnóstico e prognóstico.

Introduction: Thyroid nodules can be detected in 20%-40% of asymptomatic patients and, in 30% of them, malignant disease can be found by histopathology of post-surgical tissue samples. Attempts to establish biological links and differences between benign and malignant thyroid disorders have shown conflicting results. Previous studies using cDNA microarrays have identified genes with differential expression in thyroid diseases, including IGFBP5; proteases such as cathepsin B and proteinase inhibitors as alpha-1 antitrypsin; components involved in endocitose and intracellular transportation such as clathrin, and; cell cycle regulators such as p27 and p21. Purpose: to analyze, characterize and compare the protein expression profile of six genes differentially expressed in normal thyroid, goiter, follicular adenomas, and differentiated carcinomas, and ultimately, evaluate the clinicai importance o f the identified phenotypes. Methods: Samples of 172 tissues including 18 normal thyroids, 18 goiters, 50 follicular adenomas, 50 papillary carcinomas and 36 follicular carcinomas were collected in duplicates for analysis and comparison of six proteins expression pattems (p27, p21, clathrin, IGFBP5, alpha 1 antitrypsin and cathepsin B) using immunohistochemistry and tissue microarrays techniques. Cases were scored in relation to intensity of staining (O to 4) and number of stained cells (O to 4). Results: We found differential expression for a1-anti-tripsin, clathrin, IGFBP5, p21 and p27 when tissues were compared (p<0.0001). IGFBP5 expresswn was elevated in all neoplastic compared to non-neoplastic tissues. No significant differences were observed for cathepsin B. In normal and goiter follicular cells, immunostaining of p27 and p21 was predominantly nuclear when compared to carcinoma cells. The same pattem of expression was noted for follicular adenomas when compared to follicular carcinomas (p<0.008). Papillary carcinomas displayed a predominant diffuse cytoplasmatic p27 and p2l staining when compared to nonmalignant tissues. Conclusion: Severa! genes and new ones detected by highthroughput genomic profiling may be involved in molecular pathogenesis and tumoral progression. Transcription of these genes may result in overexpression of proteins like IGFBP5, clathrin and alpha 1 antitrypsin, while post-transcriptional changes may result in altered expression of nuclear proteins involved in cell cycle regulation that may contribute to the thyroids carcinogenesis. TMA data are consistent with results obtained with cDNA microarray showing differential protein expression for p27, p21, IGFBP5, clathrin and al-AT in normal, hyperplastic and, neoplastic thyroid tissues which could be of diagnostic and prognostic.

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