Tamoxifen inhibits transforming growth factor-alpha gene expression in human breast carcinoma samples treated with triiodothyronine

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Cell blocks allow reliable evaluation of expression of basal (CK5/6) and luminal (CK8/18) cytokeratins and smooth muscle actin (SMA) in breast carcinoma

Objective:Gene expression studies have revealed several molecular subtypes of breast carcinoma with distinct clinical and biological behaviours. DNA microarray studies correlated with immunohistochemical profiling of breast carcinomas using cytokeratin (CK) markers, Her2/neu, oestrogen receptor (ER), and basal myoepithelial cell markers have identified five breast tumour subtypes: (i) luminal A (ER+; Her2/neu-), (ii) luminal B (ER+; Her2/neu+), (iii) Her2 overexpression (ER-; Her2/neu+), (iv) basal-like (ER-; Her2/neu-, CK5/6 and 14+), and (v) negative for all markers. Luminal carcinomas express cytokeratins in a luminal pattern (CK8/18), and the basal-like type expresses CK5/6 and CK14 or basal epithelial cell markers. CK5/6, CK8/18, and smooth muscle actin (SMA) expression were assessed in cell blocks and compared with expression in surgical specimens.Methods:Sixty-two cases of breast carcinoma diagnosed by fine needle aspiration cytology with cell blocks and available surgical specimens were included. Cell blocks containing at least 10 high-power fields each with at least 10 tumour cells and surgical specimens were immunostained for CK5/6, CK8/18 and SMA.Results:Percentage sensitivity, specificity, positive predictive value, negative predictive value and accuracy were, respectively, 77, 100, 100, 92 and 94 for CK5/6; 98, 66, 96, 80 and 95 for CK8/18; and 92, 96, 85, 98 and 95 for SMA.Conclusion:The identification of CK5/6, CK8/18 and SMA by immunohistochemistry in cell blocks can be a reliable method that yields results close to those obtained in surgical specimens, and can contribute to the classification of breast carcinomas with luminal and basal expression patterns, providing helpful information in the choice of treatment and in the evaluation of prognostic and predictive factors.

Quantitative real-time RT-PCR and chromogenic in situ hybridization: precise methods to detect HER-2 status in breast carcinoma

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Detection of estrogen receptor in formalin-fixed and paraffin-embedded breast carcinoma: correlation with histological patterns.

The authors studied the expression of estrogen receptor (ER) in tissues of breast carcinomas which were previously fixed in formalin and paraffin-embedded. The ER expression was correlated with several histological findings, namely grade of differentiation, tumor necrosis, desmoplasia, lymphocytic infiltration and elastosis. The ER was detected in tissues using the avidin-biotin immunoperoxidase technique associated with the H222 monoclonal antibody from Abbott. All 39 biopsies were of infiltrating ductal carcinoma of breast and 16 of them expressed ER. The statistical analysis showed that the expression of ER was correlated with histological findings of good prognosis as well differentiated carcinomas, no tumor necrosis, absence or mild lymphocytic infiltration around the tumor cells and severe elastosis.

Shorter CAG repeat in the AR gene is associated with atypical hyperplasia and breast carcinoma

Background: Previous reports into the role of [CAG]n repeat lengths in the androgen receptor (AR) gene indicate that these may play an important part in the development and progression of breast cancer, however, knowledge regarding benign breast lesions is limited. Patients and Methods: PCR-based GeneScan analysis was used to investigate the [CAG]n repeat length at exon 1 of the AR gene in 59 benign breast lesions (27 fibroadenomas, 18 atypical hyperplasias, and 14 hyperplasias without atypia) and 54 ductal breast carcinomas. Seventy-two cancer-free women were used as a control group. In addition, [CAG]n repeats were evaluated for the presence of loss of heterozygosity (LOH) and microsatellite instability (MSI) in a subset of these samples (27 fibroadenomas, 14 hyperplasias without atypia and 22 breast carcinomas). Results: Shorter [CAG]n repeat lengths were strongly correlated with atypical hyperplasias (p=0.0209) and carcinomas (p<0.0001). LOH was found in 1/12 and 4/20 informative cases of hyperplasias without atypia and breast carcinomas, respectively. Three patients with breast carcinoma who had previously presented atypical hyperplasia showed a reduction in the [CAG]n repeat length in their carcinomas. Conclusion: Short [CAG]n repeat length (≤20) polymorphisms are strongly associated with breast carcinomas and atypical hyperplasias. Although non-significant, a subgroup of patients with breast carcinoma and genotype SS showed an association with parameters of worse outcome.

Tamoxifen inhibits transforming growth factor-α gene expression in human breast carcinoma samples treated with triiodothyronine

Objectives: To examine the effects of triiodothyronine (T3), 17β-estradiol (E2), and tamoxifen (TAM) on transforming growth factor (TGF)-α gene expression in primary breast cancer cell cultures and interactions between the different treatments. Methods and results: Patients included in the study (no.=12) had been newly diagnosed with breast cancer. Fresh human breast carcinoma tissue was cut into 0.3-mm slices. These slices were placed in six 35-mm dishes on 2-ml organ culture medium. Dishes received the following treatments: dish 1: ethanol; dish 2: T3; dish 3: T3+TAM; dish 4: TAM; dish 5: E2; dish 6: E2+TAM. TGF-α mRNA content was normalized to glyceraldehyde-3-phosphate dehydrogenase mRNA levels. All tissues included in this study were positive for estrogen receptor (ER) and thyroid hormone receptor expression. Treatment with T3 for 48 h significantly increased TGF-α mRNA levels compared to controls (15-fold), and concomitant treatment with TAM reduced expression to 3.4-fold compared to controls. When only TAM was added to the culture medium, TGF-α mRNA expression increased 5.3-fold, significantly higher than with all other treatment modalities. Conclusion: We demonstrate that TGF-α mRNA expression is more efficiently upregulated by T3 than E2. Concomitant treatment with TAM had a mitigating effect on the T3 effect, while E2 induced TGF-α upregulation. Our findings show some similarities between primary culture and breast cancer cell lines, but also some important differences: a) induction of TGF-α, a mitogenic protein, by TAM; b) a differential effect of TAM that may depend on relative expression of ER α and β; and c) supraphysiological doses of T3 may induce mitogenic signals in breast cancer tissue under conditions of low circulating E2. ©2008, Editrice Kurtis.

Fine needle aspirate cell blocks are reliable for detection of hormone receptors and HER-2 by immunohistochemistry in breast carcinoma

Aims: To evaluate the reliability of fine needle aspirate cell blocks in the assessment of oestrogen receptor (ER), progesterone receptor (PR) and HER-2/neu proteins by immunohistochemistry in comparison with surgical specimens. Materials and methods: This is a retrospective study of 62 cases of breast carcinoma diagnosed by fine needle aspiration cytology (FNAC) and confirmed using the surgical specimen. Immunohistochemical tests were performed to assess the presence of oestrogen receptor (ER), progesterone receptor (PR) and HER-2/neu proteins in cell blocks and the corresponding surgical specimens. The cell block method used alcohol prior to formalin fixation. Cases with 10% or more stained cells were considered positive for ER and PR. Positivity for HER-2/neu was assessed on a scale of 0-3+. The criterion for positivity was a score of 3+. Results: Sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and accuracy of the cell blocks in the investigation of ER, PR and HER-2/neu protein (3+) were (%): ER, 92.7, 85.7, 92.7, 85.7 and 90.3; PR, 92.7, 94.7, 97.4, 87.0 and 93.5; HER-2/neu, 70.0, 100.0, 100.0, 94.5 and 95.2. Discrepancies were seen in cell blocks in the 1+ and 2+ HER-2/neu staining scores: two of 12 cases scoring 2+ and one case of 26 scoring 1+ on cell blocks scored 3+ on surgical specimens. The correlation index between cell block and corresponding surgical specimen varied from 90% to 94%. Conclusion: Cell blocks provide a useful method of assessing ER, PR and HER-2/neu, mainly for inoperable and recurrent cases, but consideration should be given to carrying out FISH analysis on 1+ as well as 2+ HER-2/neu results. © 2012 Blackwell Publishing Ltd.

Estrogen-Responsive Genes Overlap with Triiodothyronine-Responsive Genes in a Breast Carcinoma Cell Line

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Fine needle aspiration cytology of lobular breast carcinoma and its variants

Objective: To identify associations between cytological criteria in fine needle aspiration (FNA) specimens and histological subtypes of lobular breast carcinoma (classical and other types). Study Design: FNA cytology and mastectomy specimens from 72 cases of invasive lobular breast carcinoma were consecutively retrieved from the files of the Amaral de Carvalho Hospital, Jaú-São Paulo, Brazil. All cases were reviewed regarding five cytological criteria: cellularity, cellular cohesion, presence of inflammation, nucleoli and nuclear atypia. The χ2 test or Fisher's exact tests with 95% confidence intervals (CI) were used. Results: The classical type showed lower initial cytological diagnosis of malignancy compared to the other variants (p = 0.017; odds ratio (OR) 0.26, 95% CI 0.89-0.80). Moderate/intense cellular cohesion (p = 0.011; OR 0.18, 95% CI 0.04-0.73) and mild atypia (p = 0.000; OR 16.15, 95% CI 3.20-81.48) were significantly associated with the classical type of lobular breast carcinoma, while the absence of inflammation (p = 0.082; OR 0.36, 95% CI 0.12-1.15) was marginally associated with the classical type. Conclusions: In cytology, the characterization of lobular carcinoma as malignant is difficult, especially the classical type. The association between cell cohesion and the classical type of lobular breast carcinoma may be one of the factors that complicate this diagnosis.

Imunoexpressão de receptores de estrógeno e progesterona, HER-2, P63, CK5, Catepsina D e Proteína S100A4 em carcinoma espontâneo de mama em cadelas

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Polymorphisms of Lewis and Secretor genes are related to breast cancer and metastasis in axillary lymph nodes

ABH and Lewis antigen expression has been associated with cancer development and prognosis, tumor differentiation, and metastasis. Considering that invasive ductal breast carcinoma (IDC) presents multiple molecular alterations, the aim of the present study was to determine whether the polymorphism of ABO, Lewis, and Secretor genes, as well as ABO phenotyping, could be associated with tumor differentiation and lymph nodes metastasis. Seventy-six women with IDC and 78 healthy female blood donors were submitted to ABO phenotyping/genotyping and Lewis and Secretor genotyping. Phenotyping was performed by hemagglutination and genotyping by the polymerase chain reaction with sequence-specific primers. ABO, Lewis, and Secretor genes were classified by individual single nucleotide polymorphism at sites 59, 1067, 202, and 314 of the Lewis gene, 428 of the Secretor gene, and 261 (O1 allele), 526 (O2 and B allele), and 703 (B allele). No association was found between breast cancer and ABO antigen expression (P = 0.9323) or genotype (P = 0.9356). Lewis-negative genotype was associated with IDC (P = 0.0126) but not with anatomoclinical parameters. Nonsecretor genotype was associated with axillary lymph node metastasis (P = 0.0149). In conclusion, Lewis and Secretor genotyping could be useful to predict respectively breast cancer susceptibility and axillary lymph nodes metastasis.

Estudo comparativo entre os métodos LSAB®+ e Herceptest® para a detecção de HER-2/neu em carcinoma de mama

INTRODUÇÃO: A hiperexpressão de human epidermal growth factor receptor (HER-2/neu) e a amplificação do seu gene são indicadores de formas mais agressivas do câncer de mama. A Food and Drug Administration (FDA) aprovou o teste denominado HercepTest® com a finalidade de selecionar pacientes com indicação para o uso de um anticorpo humanizado anti-HER-2/neu (trastuzumab), com efeito terapêutico comprovado. OBJETIVOS: O presente trabalho tem como objetivo comparar os resultados obtidos pelos métodos imuno-histoquímicos LSAB®+ com a utilização de anticorpo A0485 e HercepTest®. Material e métodos: Foram utilizados 50 casos de carcinoma de mama nos quais a pesquisa da hiperexpressão de HER-2 pelo método LSAB®+ já havia sido realizada. Foi repetida a pesquisa da hiperexpessão de HER-2/neu nos mesmos casos, utilizando-se o método do HercepTest®. RESULTADOS: 34 casos foram considerados negativos pelos dois métodos, com escore 0 pelo método HercepTest®. Destes, 12 obtiveram escore 1+ e 22 obtiveram escore 0 pelo método LSAB®+. em oito casos, o escore foi 2+ pelos dois métodos. Escore 3+ foi encontrado também em oito casos pelos dois métodos. DISCUSSÃO: O método mais prático utilizado em laboratório de rotina diagnóstica para investigar a hiperexpressão de HER-2/neu é o estudo imuno-histoquímico. em função de muitas variáveis, como tempo de fixação, tipo de fixador, duração da fixação, método de recuperação antigênica e tipo de anticorpo utilizado, pode haver divergência de resultados. CONCLUSÃO: O presente estudo concluiu que o método HercepTest®demonstrou resultados equivalentes aos resultados obtidos pelo método LSAB®+ em carcinoma de mama.

Avaliação da expressão da proteína bcl-2 no carcinoma de mama: estudo em punção aspirativa por agulha fina; correlação com grau histológico em espécimes cirúrgicos correspondentes

INTRODUÇÃO: O gene bcl-2 codifica uma proteína envolvida no processo de controle da apoptose. Inicialmente descrito em linfomas e posteriormente em tecidos epiteliais, sua expressão é freqüentemente encontrada em carcinomas de mama, associada a fatores de prognóstico favorável. Como a punção aspirativa por agulha fina (PAAF) tem sido utilizada como um método confiável na investigação de carcinomas de mama, acessamos a expressão de bcl-2 em material assim obtido e correlacionamos sua positividade com o grau histológico, avaliado em material cirúrgico correspondente, das respectivas pacientes, seguindo a classificação de SBR (Scarff, Bloom e Richardson). OBJETIVOS: Avaliar a expressão de bcl-2 em PAAF e correlacionar com grau histológico. METODOLOGIA: A positividade do bcl-2 foi analisada, por imunocitoquímica, em 118 casos consecutivos de PAAF e correlacionada com grau histológico em material cirúrgico correspondente, segundo classificação de SBR. RESULTADOS: A positividade para bcl-2 foi encontrada em 77 de 118 casos de PAAF (65,25%) e foi inversamente proporcional ao grau histológico (84,37%, p = 0,0022). CONCLUSÃO: A expressão de bcl-2 em PAAF correlaciona-se com fator de bom prognóstico. O índice de positividade encontrado, assim como a correlação inversa com grau histológico, está de acordo com dados publicados previamente. O fácil e rápido manejo do material obtido por PAAF permite a aplicação de técnicas complementares, de maneira confiável, como demonstra este estudo. A positividade do bcl-2 correlacionada com baixo grau histológico, assim como com outros fatores de bom prognóstico, pode, no futuro, proporcionar informação preditiva e prognóstica para pacientes candidatas a tratamento quimioterápico neo-adjuvante.