Ciclooxigenase-2 nos carcinomas ductais de mama invasivos com componente ductal in situ e no epitélio adjacente

OBJETIVO: avaliar a presença da ciclooxigenase-2 (COX-2) nos carcinomas de mama ductal in situ (CDIS) e ductal invasivo (CDI), no estroma adjacente normal e no epitélio. A correlação entre os níveis de expressão com os graus nuclear e histológico, tamanho do tumor e idade da paciente foi também analisada. MÉTODOS: foram incluídos 47 espécimes cirúrgicos provenientes de mastectomias e quadrantectomias com CDI e CDIS concomitantes, estádios clínicos I e II. Foram utilizados anticorpos policlonais anti-COX-2 para determinar a expressão da enzima. As amostras foram categorizadas em escores de zero a três, de acordo com a intensidade e o número de células coradas. RESULTADOS: COX-2 foi positivamente expressa em CDI, CDIS e epitélio normal em 86,7, 84,4 e 73,3% dos casos, respectivamente. Quanto ao grau nuclear (GN), a expressão da COX-2 foi positiva em 80% dos casos de GN-I; 81,5 e 78,9% de GN-II; e 88,5 e 96,1% de GN-III no CDIS e CDI, respectivamente. A expressão da COX-2 ocorreu em 78,9% dos CDIS com comedonecrose e em 89,3% sem comedonecrose. Quanto ao grau histológico (GH) do CDI, a COX-2 foi positiva em 83,3% de GH-I; 89,9% de GH-II e 80% de GH-III. Em relação ao diâmetro tumoral, COX-2 esteve presente em 86,1% de CDI e 83,3% de CDIS em tumores com >2 cm de diâmetro e 11% em CDI; e CDIS em tumores ≤ 2 cm. A faixa etária ≥ 50 anos apresentou 90% de expressão para CDI e 86,7% para CDIS, e 92,5% de COX-2 para ambos CDI e CDIS em pacientes com idade <50 anos. CONCLUSÕES: nossos resultados demonstram alta correlação entre as expressões da COX-2 nos CDI, CDIS e epitélio normal, o que é consistente com a hipótese de que a superexpressão da COX-2 é um evento precoce na carcinogênese mamária. Não houve diferença quando da análise do grau histológico, presença de comedonecrose, grupo etário e tamanho do tumor.

Loss of PTEN expression and AKT activation in HER2-positive breast carcinomas

PURPOSE: To examine the expression of AKT and PTEN in a series of HER2-positive primary invasive breast tumors using immunohistochemistry, and to associate these expression profiles with classic pathologic features such as tumor grade, hormone receptor expression, lymphatic vascular invasion, and proliferation.METHODS: A total of 104 HER2-positive breast carcinoma specimens were prepared in tissue microarrays blocks for immunohistochemical detection of PTEN and phosphorylated AKT (pAKT). Original histologic sections were reviewed to assess pathological features, including HER2 status and Ki-67 index values. The associations between categorical and numeric variables were identified using Pearson's chi-square test and the Mann-Whitney, respectively.RESULTS: Co-expression of pAKT and PTEN was presented in 59 (56.7%) cases. Reduced levels of PTEN expression were detected in 20 (19.2%) cases, and these 20 tumors had a lower Ki-67 index value. In contrast, tumors positive for pAKT expression [71 (68.3%)] were associated with a higher Ki-67 index value.CONCLUSION: A role for AKT in the proliferation of HER2-positive breast cancers was confirmed. However, immunohistochemical detection of PTEN expression did not correlate with an inhibition of cellular proliferation or control of AKT phosphorylation, suggesting other pathways in these mechanisms of control.

Nova classificação dos carcinomas da mama: procurando o luminal A

OBJETIVO:Comparar a distribuição das pacientes segundo os subtipos clínico-patológicos de carcinomas de mama luminais like segundo o consenso de St. Gallen 2011 e 2013.MÉTODOS:Foram selecionadas 142 pacientes com carcinoma invasivo da mama que eram positivas para receptor de estrógeno, diagnosticadas e tratadas no estado de São Paulo, região Sudeste do Brasil. A expressão dos receptores de estrógeno, progesterona (RP) e Ki-67 foi avaliada por imunoistoquímica em microarranjo de tecidos. A expressão de HER-2 foi avaliada por hibridização fluorescente in situ.RESULTADOS:Observamos que 29 casos classificados como luminais A na classificação de St. Gallen 2011 eram luminais B na classificação de 2013. Dentre os 65 casos luminais B like da classificação de 2013, além dos 29 (45%) casos que migraram, observamos 15 casos (20%) com Ki-67 >14% e pelo menos 20% das células coradas; e 21 casos (35%) com Ki-67 >14% e RP positivo em mais de 20% das células coradas.CONCLUSÕES:Comparando a distribuição das pacientes com carcinomas luminais da mama segundo a classificação de St. Gallen 2011 e 2013 observamos que houve um aumento no número de carcinomas da mama luminais B like. Consequentemente, estima-se um aumento nas indicações de quimioterapia adjuvante e no custo do tratamento.

Avaliação da superexpressão da proteína p53 e das mutações no éxon 8 do gene TP53 em carcinomas mamários caninos e glândulas normais

Este estudo foi realizado com o objetivo de avaliar a expressão da proteína p53, pela técnica de imuno-histoquímica, em neoplasmas mamários malignos em cadelas, além de investigar mutações no éxon 8 do gene supressor Tp53 por meio do padrão de bandas obtidas por PCR-RFLP. Dezenove mamas de cadelas saudáveis foram usadas como controle (Grupo 1). Amostras de 18 casos de tumores malignos (Grupo 2) e suas glândulas mamárias contralaterais (Grupo 3) foram obtidas na rotina do Hospital Veterinário da UFRPE. Os tumores foram identificados histologicamente e classificados em graus de malignidade. O método da estreptoavidina-biotina peroxidase foi utilizado para a análise da expressão de p53 por imuno-histoquímica, de acordo com a localização e intensidade da coloração. A expressão da proteína p53 não foi observada nas amostras do Grupo 1, mas foi encontrada em todas as amostras de tumores malignos (Grupo 2) seja só no núcleo, ou também no citoplasma. No Grupo 3, a expressão foi observada em quatro amostras normais e em duas que apresentavam tumor. Para a análise molecular, o DNA genômico foi extraído e submetido à PCR-RFLP com as seguintes endonucleases: AluI, BsoBI, DdeI e SmaI. O padrão de bandas foi polimórfico entre os grupos, mas não entre as variantes tumorais. Esse polimorfismo detectou mutações no fragmento estudado - éxon 8 do gene Tp53 - que podem resultar em alterações nos nucleotídeos, localizados nos sítios de restrição das enzimas. Esses achados levam a conclusão de que a imunoexpressão da p53 não tem relação com o subtipo histológico ou grau de malignidade do tumor, mas sim com a presença dos tumores no tecido mamário de cadelas. A PCR-RFLP pode ser usada como importante ferramenta para o estudo da carcinogênese mamária na cadela, possibilitando gerar diagnósticos precoces através do polimorfismo obtido com endonucleases de restrição pré-selecionadas.

Aspectos epidemiológicos, clinicopatológicos e imuno-histoquímicos de carcinomas de células escamosas vulvares em 33 vacas

Carcinomas de células escamosas vulvares (CCEVs) em bovinos foram estudados retrospectivamente quanto à prevalência, epidemiologia, quadro clinicopatológico e aspectos imuno-histoquímicos. O grau de pigmentação da pele vulvar foi também avaliado. Nos 48 anos analisados retrospectivamente, foram computados materiais de necropsias e biópsias de 7.483 bovinos recebidos no Laboratório de Patologia Veterinária da UFSM. Desses, em 664 (8,87%) casos de neoplasmas foram identificados, sendo 33 (4,97%) casos de CCEVs. Dezenove eram vacas da raça Holandesa, três da Charolesa, uma era Jersey e 10 eram sem raça definida. A principal alteração macroscópica foi aumento de volume vulvar, sangrante e com miíase concomitante. As massas tumorais eram firmes, ulceradas e com áreas amarelas. Foi possível reavaliar microscopicamente 30 dos 33 casos. Desses, oito eram CCEVs bem diferenciados, 17 eram moderadamente diferenciados e cinco eram pobremente diferenciados. A avaliação de lesões intraepiteliais escamosas (LIEs) foi realizada em tecidos de 21 casos que tinham epitélio de revestimento. Hiperplasia epitelial foi observada em 10 casos; displasia leve em dois, moderada em um e acentuada em cinco casos; em três casos não havia LIEs. A técnica de Fontana-Masson para melanina foi realizada em 21 casos. Desses, em 17 a pigmentação do epitélio da epiderme vulvar era ausente, em dois era leve e em outros dois era moderada. Independentemente do grau de diferenciação dos CCEVs, houve imunomarcação acentuada da maioria dos ceratinócitos neoplásicos para pancitoceratina bovina pela técnica de imuno-histoquímica (IHQ). Papilomavírus bovino não foi detectado pela IHQ neste estudo.

Avaliação da imunomarcação de células-tronco tumorais em carcinossarcomas mamários e carcinomas em tumores mistos em cadelas

As células-tronco tumorais (CTTs) pertencem a uma pequena população de células dentro do tumor com propriedades de autorrenovação e diferenciação em outros tipos celulares. Neste estudo avaliou-se o comportamento tanto das porções mesenquimais quanto das epiteliais de seis carcinossarcomas (CSs), 11 carcinomas em tumores mistos (CTMs) grau I, 11 grau II e 10 grau III. Nas porções epiteliais dos CS e CTM foram observadas imunomarcações para os anticorpos CD44, CD24, Oct-4 e ALDH-1. Nas porções mesenquimais dos CS, nas porções epiteliais dos CTMs graus II e III não houve imunomarcação para o ALDH-1. Concluiu-se que as CTTs são expressas em proporções iguais tanto nas porções mesenquimais quanto nas epiteliais dos CSs e ausentes nas porções mesenquimais bem diferenciadas de CTMs.

Immunodetection of myeloid and plasmacytoid dendritic cells in mammary carcinomas of female dogs

ABSTRACT: Dendritic cells have attracted great interest from researchers as they may be used as targets of tumor immune evasion mechanisms. The main objective of this study was to evaluate the relationship between the dendritic cells (DCs) subpopulation in simple type mammary carcinomas in female dogs. Two groups of samples were used: the control group consisted of 18 samples of mammary tissue without changes and the tumor group with 26 simple type mammary carcinomas. In these groups, we evaluated the immunodetection of immature and mature myeloid DCs, plasmacytoid DCs and MHC-II. In mammary tumor, mature myeloid DCs predominated in the peritumoral region, while immature myeloid DCs and plasmacytoid DCs were evident in the intratumoral region. Immunostaining of MHC-II was visualized in mammary acini (control group), in tumor cells and inflammatory infiltration associated with tumors. The comparison between the control and tumor groups showed a statistically significant difference between immature myeloid DCs, mature myeloid DCs and plasmacytoid DCs. The immunodetection of MHC-II was not significant when comparing the groups. The predominance of immature DCs in the tumor group is possibly related to an inefficient immune response, promoting the development and survival of tumor cells. The presence of plasmacytoid DCs in the same group suggests a worse prognosis for female dogs with mammary tumors. Therefore, the ability of differentiation of canine dendritic cells could be influenced by neoplastic cells and by the tumor microenvironment.

Genetic alterations in head and neck squamous cell carcinomas

The genetic alterations observed in head and neck cancer are mainly due to oncogene activation (gain of function mutations) and tumor suppressor gene inactivation (loss of function mutations), leading to deregulation of cell proliferation and death. These genetic alterations include gene amplification and overexpression of oncogenes such as myc, erbB-2, EGFR and cyclinD1 and mutations, deletions and hypermethylation leading to p16 and TP53 tumor suppressor gene inactivation. In addition, loss of heterozygosity in several chromosomal regions is frequently observed, suggesting that other tumor suppressor genes not yet identified could be involved in the tumorigenic process of head and neck cancers. The exact temporal sequence of the genetic alterations during head and neck squamous cell carcinoma (HNSCC) development and progression has not yet been defined and their diagnostic or prognostic significance is controversial. Advances in the understanding of the molecular basis of head and neck cancer should help in the identification of new markers that could be used for the diagnosis, prognosis and treatment of the disease.

Biosynthesis and metabolism of steroid hormones by human adrenal carcinomas

Over a 15-year period, our university-based laboratory obtained 125 adrenal tumors, of which 15 (12%) were adrenal cortical carcinomas. Of these, 6 (40% of the carcinomas) occurred in patients with clear clinical manifestations of steroid hormone excess. Adrenal cortical carcinoma cells derived from the surgically resected tumors in 4 of these patients were isolated and established in primary culture. Radiotracer steroid interconversion studies were carried out with these cultures and also on mitochondria isolated from homogenized tissues. Large tumors had the lowest steroidogenic activities per weight, whereas small tumors had more moderately depressed enzyme activities relative to cells from normal glands. In incubations with pregnenolone as substrate, 1 mM metyrapone blocked the synthesis of corticosterone and cortisol and also the formation of aldosterone. Metyrapone inhibition was associated with a concomitant increase in the formation of androgens (androstenedione and testosterone) from pregnenolone. Administration of metyrapone in vivo before surgery in one patient resulted in a similar increase in plasma androstenedione, though plasma testosterone levels were not significantly affected. In cultures of two of four tumors examined, dibutyryl cAMP stimulated 11ß-hydroxylase activity modestly; ACTH also had a significant stimulatory effect in one of these tumors. Unlike results obtained with normal or adenomatous adrenal cortical tissues, mitochondria from carcinomatous cells showed a lack of support of either cholesterol side-chain cleavage enzyme complex or steroid 11ß-hydroxylase activity by Krebs cycle intermediates (10 mM isocitrate, succinate or malate). This finding is consistent with the concept that these carcinomas may tend to function predominantly in an anaerobic manner, rather than through the oxidation of Krebs cycle intermediates.

Low frequency of p53 mutations in cervical carcinomas among Brazilian women

Human papillomavirus (HPV) infections of the high-risk types are strongly linked to the development of cervical carcinoma. The HPV oncoproteins E6 and E7 are thought to play a crucial role in this process through their interactions with the p53 protein and the retinoblastoma susceptibility gene product pRb, respectively. E6 binds to p53 protein promoting its degradation. This is considered to contribute to the oncogenesis of HPV-associated anogenital cancer. On the other hand, in HPV-negative cervical carcinoma, p53 mutations are thought to have a role in the transformation process. A total of 122 HPV-positive cervical carcinoma tissue samples were evaluated for the presence of mutations in exons 5-8 of the p53 gene by single-stranded conformation polymorphism analysis and DNA sequencing. Only four missense point mutations were detected. These findings suggest that other mechanisms independent of p53 inactivation may play a role in the genesis of cervical carcinomas.

Does the correlation between EBNA-1 and p63 expression in breast carcinomas provide a clue to tumorigenesis in Epstein-Barr virus-related breast malignancies?

Several investigators have identified Epstein-Barr virus (EBV) particles in breast carcinomas, a fact that supports a role for EBV in mammary tumorigenesis. The possible mechanism involved in this process is not clear. The present study was carried out in an attempt to determine whether there is a relationship between latent infection with EBV and p53 and p63 expression in breast carcinomas. Immunohistochemistry developed with 3.3-diaminobenzidine tetrahydrochloride was performed in 85 formalin-fixed paraffin-embedded breast carcinomas using anti-EBV EBNA-1, anti-p63, anti-p53, anti-estrogen receptor (ER) and anti-progesterone receptor (PR) antibodies. The cases were selected to represent each of the various histologic types: intraductal carcinoma (N = 12), grade I invasive ductal carcinoma (N = 15), grade II invasive ductal carcinoma (N = 15), grade III invasive ductal carcinoma (N = 15), tubular carcinoma (N = 8), lobular carcinoma (N = 10), and medullary carcinoma (N = 10). The ductal breast carcinomas were graded I, II and III based on the Scarff-Bloom and Richardson grading system modified by Elston and Ellis. One slide containing at least 1000 neoplastic cells was examined in each case. ER, PR, p63, p53 and EBNA-1 were positive in 60, 40, 11.8, 21.2 and 37.6% of carcinomas, respectively. There was a correlation between EBNA-1 and p63 expression (P < 0.001), but not between EBNA-1 and p53 (P = 0.10). These data suggest a possible role for p63 in the mammary tumorigenesis associated with Epstein-Barr virus infection.

p53 immunostaining is correlated with reduced survival and is not correlated with gene mutations in resected pulmonary large cell carcinomas

Malignancy of pulmonary large cell carcinomas (LCC) increases from classic LCC through LCC with neuroendocrine morphology (LCCNM) to large cell neuroendocrine carcinomas (LCNEC). However, the histological classification has sometimes proved to be difficult. Because the malignancy of LCC is highly dependent on proteins with functions in the cell cycle, DNA repair, and apoptosis, p53 has been targeted as a potentially useful biological marker. p53 mutations in lung cancers have been shown to result in expression and protein expression also occurs in the absence of mutations. To validate the importance of both p53 protein expression (by immunostaining) and p53 gene mutations in lung LCC (by PCR-single strand conformational polymorphism analysis of exons 5, 6, 7, and 8) and to study their relationships with clinical factors and sub-classification we investigated the correlation of p53 abnormalities in 15 patients with LCC (5 classic LCC, 5 LCNEC, and 5 LCCNM) who had undergone resection with curative intent. Of these patients, 5/15 expressed p53 and none had mutant p53 sequences. There was a negative survival correlation with positive p53 immunostaining (P = 0.05). After adjustment for stage, age, gender, chemotherapy, radiotherapy, and histological subtypes by multivariate analysis, p53 expression had an independent impact on survival. The present study indicates that p53 assessment may provide an objective marker for the prognosis of LCC irrespective of morphological variants and suggests that p53 expression is important for outcome prediction in patients with the early stages of LCC. The results reported here should be considered to be initial results because tumors from only 15 patients were studied: 5 each from LCC, LCNEC and LCCNM. This was due to the rarity of these specific diseases.

Differential gene expression profiles of hepatocellular carcinomas associated or not with viral infection

Chronic hepatitis B (HBV) and C (HCV) virus infections are the most important factors associated with hepatocellular carcinoma (HCC), but tumor prognosis remains poor due to the lack of diagnostic biomarkers. In order to identify novel diagnostic markers and therapeutic targets, the gene expression profile associated with viral and non-viral HCC was assessed in 9 tumor samples by oligo-microarrays. The differentially expressed genes were examined using a z-score and KEGG pathway for the search of ontological biological processes. We selected a non-redundant set of 15 genes with the lowest P value for clustering samples into three groups using the non-supervised algorithm k-means. Fisher’s linear discriminant analysis was then applied in an exhaustive search of trios of genes that could be used to build classifiers for class distinction. Different transcriptional levels of genes were identified in HCC of different etiologies and from different HCC samples. When comparing HBV-HCC vs HCV-HCC, HBV-HCC/HCV-HCC vs non-viral (NV)-HCC, HBC-HCC vs NV-HCC, and HCV-HCC vs NV-HCC of the 58 non-redundant differentially expressed genes, only 6 genes (IKBKβ, CREBBP, WNT10B, PRDX6, ITGAV, and IFNAR1) were found to be associated with hepatic carcinogenesis. By combining trios, classifiers could be generated, which correctly classified 100% of the samples. This expression profiling may provide a useful tool for research into the pathophysiology of HCC. A detailed understanding of how these distinct genes are involved in molecular pathways is of fundamental importance to the development of effective HCC chemoprevention and treatment.

Expression of E-cadherin, Snail and Hakai in epithelial cells isolated from the primary tumor and from peritumoral tissue of invasive ductal breast carcinomas

Epithelial intercellular cohesion, mainly mediated by E-cadherin (CDH1) expression and function, may be deregulated during cancer cell invasion of adjacent tissues and lymphatic and vascular channels. CDH1 expression is down-modulated in invasive lobular breast carcinomas but its regulation in invasive ductal carcinomas (IDC) is less clear. CDH1 expression is repressed by transcription factors such as Snail (SNAI1) and its product is degraded after Hakai ubiquitination. We compared CDH1, SNAI1 and HAKAI mRNA expression in IDC and paired adjacent normal breast tissue and evaluated its relation with node metastasis and circulating tumor cells. Matched tumor/peritumoral and blood samples were collected from 30 patients with early IDC. Epithelial cells from each compartment (tumor/peritumoral) were recovered by an immunomagnetic method and gene expression was determined by real time RT-PCR. There were no differences in CDH1, SNAI1 and HAKAI mRNA expression between tumor and corresponding peritumoral samples and no differential tumoral gene expression according to nodal involvement. Another 30 patients with a long-term follow-up (at least 5 years) and a differential prognosis (good or poor, as defined by breast cancer death) had E-cadherin and Snail protein detected by immunohistochemistry in tumor samples. In this group, E-cadherin-positive expression, but not Snail, may be associated with a better prognosis. This is the first report simultaneously analyzing CDH1, SNAI1 and HAKAI mRNA expression in matched tumor and peritumoral samples from patients with IDC. However, no clear pattern of their expression could distinguish the invasive tumor compartment from its adjacent normal tissue.