Comparative genomic hybridization analysis of benign and invasive male breast neoplasms

Comparative genomic hybridization (CGH) analysis was performed for the identification of chromosomal imbalances in two benign gynecomastias and one malignant breast carcinoma derived from patients with male breast disease and compared with cytogenetic analysis in two of the three cases. CGH analysis demonstrated overrepresentation of 8q in all three cases. One case of gynecomastia presented gain of 1p34.3similar topter. 11p14similar toq12. and 17p11.2similar toqter, and loss of 1q41similar toqter and 4q33similar toqter. The other gynecomastia presented del(1)(q41) as detected by both cytogenetic and CGH analysis. CGH analysis of the invasive ductal carcinoma confirmed a gain of 17p11.2similar toqter previously detected by cytogenetic analysis. These regions showed some similarity in their pattern of imbalance to the chromosomal alterations described in female and male breast cancer. (C) 2002 Elsevier B.V. All rights reserved.

CDHI promoter hypermethylation and E-cadherin protein expression in infiltrating breast cancer

Background: the E-cadherin gene (CDH1) maps, at chromosome 16q22.1, a region often associated with loss of heterozygosity (LOH) in human breast cancer. LOH at this site is thought to lead to loss of function of this tumor suppressor gene and was correlated with decreased disease-free survival, poor prognosis, and metastasis. Differential CpG island methylation in the promoter region of the CDH1 gene might be an alternative way for the loss of expression and function of E-cadherin, leading to loss of tissue integrity, an essential step in tumor progression.Methods: the aim of our study was to assess, by Methylation-Specific Polymerase Chain Reaction (MSP), the methylation pattern of the CDH1 gene and its possible correlation with the expression of E-cadherin and other standard immunohistochemical parameters (Her-2, ER, PgR, p53, and K-67) in a series of 79 primary breast cancers ( 71 infiltrating ductal, 5 infiltrating lobular, 1 metaplastic, 1 apocrine, and 1 papillary carcinoma).Results: CDH1 hypermethylation was observed in 72% of the cases including 52/71 ductal, 4/5 lobular carcinomas and 1 apocrine carcinoma. Reduced levels of E-cadherin protein were observed in 85% of our samples. Although not statistically significant, the levels of E-cadherin expression tended to diminish with the CDH1 promoter region methylation. In the group of 71 ductal cancinomas, most of the cases of showing CDH1 hypermethylation also presented reduced levels of expression of ER and PgR proteins, and a possible association was observed between CDH1 methylation and ER expression ( p = 0.0301, Fisher's exact test). However, this finding was not considered significant after Bonferroni correction of p-value.Conclusion: Our preliminary findings suggested that abnormal CDH1 methylation occurs in high frequencies in infiltrating breast cancers associated with a decrease in E-cadherin expression in a subgroup of cases characterized by loss of expression of other important genes to the mammary carcinogenesis process, probably due to the disruption of the mechanism of maintenance of DNA methylation in tumoral cells.

Absence of transforming growth factor-beta type II receptor is associated with poorer prognosis in HER2-negative breast tumours

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

Absence of TGF-beta RII predicts bone and lung metastasis and is associated with poor prognosis in stage III breast tumors

In the case of operated breast cancer (BC), prognostic markers help to determine if the patient needs additional treatment and predictive markers help the clinician to decide which treatment to use. Thus, a better knowledge of known predictive and prognostic markers and the identification of new markers, may improve the treatment of BC patients. The transforming growth factor-beta type II receptor (TGF-beta RII), a main receptor of transforming growth factor beta pathway, is a potential new prognostic marker. The aims of the present study were to investigate both the predictive and prognostic impact of TGF-beta RII in BC samples. TGF-beta RII protein expression was evaluated using immunohistochemistry on a tissue microarray containing 110 TNM stage III BC samples obtained prior to doxorubicin-based neoadjuvant chemotherapy (NAC). Our results demonstrate that TGF-beta RII did not predict the response to NAC. on the other hand, an association between TGF-beta RII-negative tumor and higher risk of metastasis to lungs and bones was verified. TGF-beta RII negativity was an independent prognostic factor for decreased disease-free and overall survival.

Association of matrix metalloproteinase-8 gene variation with breast cancer prognosis

Animal and cell studies indicate an inhibitory effect of matrix metalloproteinase-8 (MMP8) on tumorigenesis and metastasis. We investigated whether MMP8 gene variation was associated with breast cancer metastasis and prognosis in humans. We first studied nine tagging single nucleotide polymorphisms (SNP) in the MMP8 gene in 140 clinically and pathologically well-characterized breast cancer patients. Four of the SNPs were found to be associated with lymph node metastasis, the most pronounced being a promoter SNP (rs11225395) with its minor allele (T) associating with reduced susceptibility to lymph node metastasis (P = 0.02). This SNP was further evaluated for association with cancer relapse and survival among a cohort of similar to 1,100 breast cancer patients who had been followed for cancer recurrence and mortality for a median of 7.1 years. The T allele was associated with reduced cancer relapse and greater survival, particularly among patients with earlier stage cancer. Among patients of tumor-node-metastasis stage 0 to 11, the adjusted hazard ratio of disease-free survival was 0.7 [95% confidence interval (95% CI), 0.5-0.9] for patients carrying T allele compared with those homozygous for the C allele (P = 0.02). In vitro experiments showed that the T allele had higher promoter activity than the C allele in breast cancer cells. Electrophoretic mobility shift assays showed binding of nuclear proteins to the DNA sequence at the SNP site of the T allele but not that of the C allele. The data suggest that MMP8 gene variation may influence breast cancer prognosis and support the notion that MMP8 has an inhibitory effect on cancer metastasis.

Profile of thyroid hormones in breast cancer patients

Estrogen involvement in breast cancer has been established; however, the association between breast cancer and thyroid diseases is controversial. Estrogen-like effects of thyroid hormone on breast cancer cell growth in culture have been reported. The objective of the present study was to determine the profile of thyroid hormones in breast cancer patients. Serum aliquots from 26 patients with breast cancer ranging in age from 30 to 85 years and age-matched normal controls (N = 22) were analyzed for free triiodothyronine (T3F), free thyroxine (T4F), thyroid-stimulating hormone (TSH), antiperoxidase antibody (TPO), and estradiol (E2). Estrogen receptor ß (ERß) was determined in tumor tissues by immunohistochemistry. Thyroid disease incidence was higher in patients than in controls (58 vs 18%, P < 0.05). Subclinical hyperthyroidism was the most frequent disorder in patients (31%); hypothyroidism (8%) and positive anti-TPO antibodies (19%) were also found. Subclinical hypothyroidism was the only dysfunction (18%) found in controls. Hyperthyroidism was associated with postmenopausal patients, as shown by significantly higher mean T3 and T4 values and lower TSH levels in this group of breast cancer patients than in controls. The majority of positive ERß tumors were clustered in the postmenopausal patients and all cases presenting subclinical hyperthyroidism in this subgroup concomitantly exhibited Erß-positive tumors. Subclinical hyperthyroidism was present in only one of 6 premenopausal patients. We show here that postmenopausal breast cancer patients have a significantly increased thyroid hormone/E2 ratio (P < 0.05), suggesting a possible tumor growth-promoting effect caused by this misbalance.

P53 PROTEIN EXPRESSION AND NUCLEAR-DNA CONTENT IN BREAST INTRADUCTAL PROLIFERATIONS

Immunohistochemical analysis of the p53 gene protein and cytometric assessment of nuclear DNA were performed in a series of 51 cases of intraductal breast proliferation. The series included 22 cases of intraductal hyperplasia without atypia, 6 cases of intraductal hyperplasia with atypia, and 23 cases of pure intraductal carcinoma. Expression of p53 protein was detected in one case of intraductal hyperplasia without atypia (4.5 per cent), one case of intraductal hyperplasia with atypia (16.6 per cent) and six cases of intraductal carcinoma (26.0 per cent). No significant correlation was observed between p53 expression and histological subtype of intraductal carcinoma. Aneuploidy was demonstrated in two cases of intraductal hyperplasia with atypia (33.3 per cent) and in 18 cases of intraductal carcinoma (78.2 per cent). All cases of intraductal hyperplasia without atypia were euploid. No significant association was observed between p53 protein expression and ploidy in intraductal hyperplasia. The only case of intraductal hyperplasia without atypia positive for p53 was euploid, whereas the only p53-positive case of intraductal hyperplasia with atypia was aneuploid. Among the intraductal carcinomas, only the aneuploid cases showed positivity for p53, regardless of histological subtype. The results suggest that some of the changes observed in invasive breast carcinoma, such as p53 expression and aneuploidy, are already present in breast intraductal proliferation, especially in areas with atypia and in intraductal carcinoma. The expression of p53 in breast intraductal proliferation may reflect the acquisition of p53 gene mutations in cells unable adequately to repair DNA damage, with genomic instability which would lead to clonal expansion and putative evolution to invasive disease.

Simple mucin-type carbohydrate antigens (T, sialosyl T, Tn and sialosyl-Tn) in breast carcinogenesis

Immunohistochemical analysis of the expression of simple mucin-type carbohydrate antigens (Tn, sialyl-Tn and T) was performed in a series of 43 cases of intraductal hyperplasia without atypia, 9 cases of intraductal hyperplasia with atypia, 54 cases of ductal carcinoma in situ (DCIS) and 26 cases of invasive breast carcinoma. We also studied 36 cases of isolated breast normal epithelium, 20 cases of 'normal' breast epithelium adjacent to neoplasms and 14 cases of apocrine metaplasia. All antigens were detected in different frequencies in normal, hyperplastic, metaplastic and neoplastic breast epithelium. Tn and sialyl-Tn are expressed more frequently in malignant than in benign breast epithelium; while Tn expression increases from normal to invasive carcinomas, sialyl-Tn increases until DCIS and drops in invasive carcinomas, suggesting that either there is a failure of a proportion of DCIS to progress to invasive carcinoma or loss of expression of sialyl-Tn when some carcinomas become invasive. The high frequency of Tn and sialyl-Tn expression in breast intraductal proliferations probably reflects incomplete glycosylation in these lesions, which is a well-known tumour-associated phenomenon and supports the assumption that such lesions are putative precursors of breast cancer. T antigen was expressed in all groups studied, but its prevalence differed significantly between normal and neoplastic epithelium. The expression of these antigens in epithelium adjacent to carcinomas is similar to that found in isolated normal breast epithelium, whereas apocrine metaplasia has a pattern of simple mucin-type glycosylation that is specific and distinct from that of the normal breast epithelium, with a high frequency of marked expression of Tn and sialyl-Tn. The similarity of the pattern of expression of simple mucin-type antigens in metaplasia and malignant neoplasia reduces the usefulness of these markers from a diagnostic standpoint.

Tamoxifen down-regulates CaMKII messenger RNA levels in normal human breast tissue

Tamoxifen was proven to reduce the incidence of breast cancer by 49% in women at increased risk of the disease in the Breast Cancer Prevention Trial. In order to identify potential candidates to explain the preventive effect induced by tamoxifen on breast cancer, normal breast tissue obtained from 42 fibroadenoma patients, randomly assigned to receive placebo or tamoxifen, was analyzed by the reverse Northern blot and RT-PCR techniques. The cDNA fragments used on Northern blot membranes were generated by the Human Cancer Genome Project funded by the Ludwig Institute for Cancer Research and FAPESP (Fundação de Amparo à Pesquisa do Estado de São Paulo, Brazil). Total RNA was obtained from normal breast tissue from patients with clinical, cytological and ultrasound diagnosis of fibroadenoma. After a 50-day treatment with tamoxifen (10 or 20 mg/day) or placebo, normal breast tissue adjacent to the tumor was collected during lumpectomy with local anesthesia. One differentially expressed gene, Calcium/calmodulin-dependent protein kinase II (CaMKII), was found to be down-regulated during TAM treatment. CaMKII is an ubiquitous serine/threonine protein kinase that has been implicated in the diverse effects of hormones utilizing Ca2+ as a second messenger as well as in c-fos activation. These results indicate that the down-regulation of CaMKII induced by TAM might represent alternative or additional mechanisms of the action of this drug on cell cycle control and response to hormones in normal human breast tissue.

Adjuvant therapy with GnRH agonists/tamoxifen in breast cancer should be a good council for patients with hormone receptor-positive tumours and wish to preserve fertility

Infertility represents one of the main long-term consequences of combination chemotherapy used for the treatment of breast cancer. Approximately 60%-65% of breast cancers express the nuclear hormone receptor in premenopausal women. Adjuvant endocrine therapy is an integral component of care for patients with hormone receptor-positive (HR+) tumours. The GnRH agonist (GnRHa) alone or in combination with tamoxifen produces results at least similar to those obtained with the different chemotherapy protocols in patients with HR+ tumors with respect to recurrence-free survival and overall survival, Presentation of the hypothesis: It is time to indicate adjuvant therapy with GnRHa associated with tamoxifen for patients with breast cancer (HR+ tumours) if they want to preserve their reproductive function. Testing the hypothesis: Assessment of ovarian reserve tests: follicle stimulating hormone (FSH), anti-Mullerian hormone (AMH), inhibin B, antral follicle count (AFC) and ovarian volume 6 months, and 1 year after the end of therapy with GnRHa/tamoxifen. The recurrence-free survival and overall survival should be analysed. Implications of the hypothesis: The major implication will be to avoid adjuvant chemotherapy for patients with breast cancer (HR+ tumours) that request fertility preservation. It is expected that ovarian function should not be altered in almost all cases. © Todos os direitos reservados a SBRA - Sociedade Brasileira de Reprodução Assistida.

Necrotizing soft tissue infection of the breast: Case report and literature review

Background: Necrotizing soft tissue infection (NSTI) is characterized by progressive infectious gangrene of the skin and subcutaneous tissue. Its treatment involves intensive care, broad-spectrum antibiotic therapy, and full debridement. Methods: We present two cases of NSTI of the breast, adding these cases to the 14 described in the literature, reviewing the characteristics and evolution of all cases. Case Report: On the fourth day after mastectomy, a 59-year-old woman with ulcerated breast cancer developed Type I NSTI caused by Pseudomonas aeruginosa, which had a favorable evolution after debridement and broad-spectrum antibiotics. The second patient was a 57-year-old woman submitted to a mastectomy and axillary dissection, who had recurrent seromas. On the 32nd post-operative day, after a seroma puncture, she developed Type II NSTI caused by β-hemolytic streptococci. She developed sepsis and died on the tenth day after debridement, intensive care, and broad-spectrum antibiotics. The cases are the first description of breast NSTI after mammary seroma aspiration and the first report of this condition caused by P. aeruginosa. Conclusion: Necrotizing soft tissue infection is rare in breast tissue. It frequently is of Type II, occurring mainly after procedures in patients with breast cancer. The surgeon's participation in controlling the focus of the infection is of fundamental importance, and just as important are broad-spectrum antibiotic therapy and support measures, such as maintenance of volume, correction of electrolytic disorders, and treatment of sepsis and septic shock. Once the infection has been brought under control, skin grafting or soft tissue flaps can be considered. The mortality rate in breast NSTI is 18.7%, all deaths being in patients with the fulminant Type II form. Surgical oncologists need to be alert to the possibility of this rare condition. © 2012, Mary Ann Liebert, Inc.

Pain and quality of life in patients undergoing radiotherapy for spinal metastatic disease treatment

Background: Radiotherapy is an important tool in the control of pain in patients with spinal metastatic disease. We aimed to evaluate pain and of quality of life of patients with spinal metastatic disease undergoing radiotherapy with supportive treatment. Methods. The study enrolled 30 patients. From January 2008 to January 2010, patients selection included those treated with a 20Gy tumour dose in five fractions. Patients completed the visual analogue scale for pain assessment and the SF-36 questionnaire for quality of life assessment. Results: The most frequent primary sites were breast, multiple myeloma, prostate and lymphoma. It was found that 14 spinal metastatic disease patients (46.66%) had restricted involvement of three or fewer vertebrae, while 16 patients (53.33%) had cases involving more than three vertebrae. The data from the visual analogue scale evaluation of pain showed that the average initial score was 5.7 points, the value 30days after the end of radiotherapy was 4.60 points and the average value 6months after treatment was 4.25 points. Notably, this final value was 25.43% lower than the value from the initial analysis. With regard to the quality of life evaluation, only the values for the functional capability and social aspects categories of the questionnaire showed significant improvement. Conclusion: Radiotherapy with supportive treatment appears to be an important tool for the treatment of pain in patients with spinal metastatic disease. © 2013 Valesin Filho et al; licensee BioMed Central Ltd.

Risk of metabolic syndrome in postmenopausal breast cancer survivors

The aim of this study was to assess the risk of metabolic syndrome (MetS) in postmenopausal breast cancer survivors as compared with postmenopausal women without breast cancer. METHODS: In this cross-sectional study, 104 postmenopausal breast cancer survivors were compared with 208 postmenopausal women (controls) attending a university hospital. Eligibility criteria included the following: amenorrhea longer than 12 months and aged 45 years or older, treated for breast cancer, and metastasis-free for at least 5 years. The control group consisted of women with amenorrhea longer than 12 months and aged 45 years or older and without breast cancer, matched by age and menopause status (in a proportion of 1:2 as sample calculation). Clinical and anthropometric data were collected. Biochemical parameters, including total cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, triglycerides, glucose, and C-reactive protein, were measured. Women showing three or more diagnostic criteria were diagnosed as having MetS: waist circumference of 88 cm or larger, blood pressure of 130/85 mm Hg or higher, triglycerides level of 150 mg/dL or higher, high-density lipoprotein cholesterol level lower than 50 mg/dL, and glucose level of 100 mg/dL or higher. For statistical analysis, Student's t test, χ2 test, and logistic regression (odds ratio [OR]) were used. RESULTS: The mean (SD) age of breast cancer survivors was 60.6 (8.6) years, with a mean (SD) follow-up of 9.4 (4.4) years. A higher percentage of breast cancer survivors (46.2%) were obese as compared with controls (32.7%; P < 0.05), and a smaller percentage showed optimal values for low-density lipoprotein cholesterol, glucose, and C-reactive protein versus controls (P < 0.05). MetS was diagnosed in 50% of breast cancer survivors and in 37.5% of control group women (P < 0.05). Among the MetS diagnostic criteria, the most prevalent was abdominal obesity (waist circumference >88 cm), affecting 62.5% and 67.8% of the participants, respectively. In the control group, breast cancer survivors had a higher risk for MetS (OR, 1.66; 95% CI, 1.04-2.68), dysglycemia (OR, 1.05; 95% CI, 1.09-3.03), and hypertension (OR, 1.71; 95% CI, 1.02-2.89). CONCLUSIONS: Postmenopausal breast cancer survivors present a higher risk of developing MetS as compared with women without breast cancer. © 2012 by The North American Menopause Society.

N-acetylglucosaminidase, myeloperoxidase and vascular endothelial growth factor serum levels in breast cancer patients

Inflammatory cells surround breast carcinomas and may act promoting tumor development or stimulating anti-tumor immunity. N-acetylglucosaminidase (NAG) has been employed to detect macrophage accumulation/activation. Myeloperoxidase (MPO) is considered a marker for neutrophils activity/accumulation. Vascular Endothelial Growth Factor (VEGF) is as strong pro-angiogenic cytokine. The aim of this study was to measure the systemic inflammatory response by measuring serum levels of NAG, MPO and VEGF in women diagnosed with breast cancer and associate this response to the peritumoral inflammatory infiltrate and to prognostic factors. Serum samples obtained from women with no evidence of disease (n = 31) and with breast cancer (n = 68) were analyzed for the activities of NAG, MPO and VEGF by enzymatic assay. Serum levels of NAG and VEGF were higher in healthy volunteers (P < 0.0001) and serum levels of MPO were higher in patients with breast cancer (P = 0.002). Serum levels of NAG were positively correlated to serum levels of MPO and VEGF (P < 0.0001 and P = 0.0012, respectively) and MPO and VEGF serum levels had also a positive correlation (P = 0.0018). The inflammatory infiltrate was not associated to serum levels of the inflammatory markers, and higher levels of MPO were associated to lymphovascular invasion negativity (P = 0.0175). (C) 2013 Elsevier Masson SAS. All rights reserved.

ATM down-regulation is associated with poor prognosis in sporadic breast carcinomas

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