Circulating sex hormones and breast cancer risk factors in postmenopausal women: reanalysis of 13 studies.

BACKGROUND: Breast cancer risk for postmenopausal women is positively associated with circulating concentrations of oestrogens and androgens, but the determinants of these hormones are not well understood. METHODS: Cross-sectional analyses of breast cancer risk factors and circulating hormone concentrations in more than 6000 postmenopausal women controls in 13 prospective studies. RESULTS: Concentrations of all hormones were lower in older than younger women, with the largest difference for dehydroepiandrosterone sulphate (DHEAS), whereas sex hormone-binding globulin (SHBG) was higher in the older women. Androgens were lower in women with bilateral ovariectomy than in naturally postmenopausal women, with the largest difference for free testosterone. All hormones were higher in obese than lean women, with the largest difference for free oestradiol, whereas SHBG was lower in obese women. Smokers of 15+ cigarettes per day had higher levels of all hormones than non-smokers, with the largest difference for testosterone. Drinkers of 20+ g alcohol per day had higher levels of all hormones, but lower SHBG, than non-drinkers, with the largest difference for DHEAS. Hormone concentrations were not strongly related to age at menarche, parity, age at first full-term pregnancy or family history of breast cancer. CONCLUSION: Sex hormone concentrations were strongly associated with several established or suspected risk factors for breast cancer, and may mediate the effects of these factors on breast cancer risk.

Bone mineral density in breast cancer patients treated with adjuvant letrozole, tamoxifen, or sequences of letrozole and tamoxifen in the BIG 1-98 study (SAKK 21/07).

BACKGROUND: The risk of osteoporosis and fracture influences the selection of adjuvant endocrine therapy. We analyzed bone mineral density (BMD) in Swiss patients of the Breast International Group (BIG) 1-98 trial [treatment arms: A, tamoxifen (T) for 5 years; B, letrozole (L) for 5 years; C, 2 years of T followed by 3 years of L; D, 2 years of L followed by 3 years of T]. PATIENTS AND METHODS: Dual-energy X-ray absorptiometry (DXA) results were retrospectively collected. Patients without DXA served as control group. Repeated measures models using covariance structures allowing for different times between DXA were used to estimate changes in BMD. Prospectively defined covariates were considered as fixed effects in the multivariable models. RESULTS: Two hundred and sixty-one of 546 patients had one or more DXA with 577 lumbar and 550 hip measurements. Weight, height, prior hormone replacement therapy, and hysterectomy were positively correlated with BMD; the correlation was negative for letrozole arms (B/C/D versus A), known osteoporosis, time on trial, age, chemotherapy, and smoking. Treatment did not influence the occurrence of osteoporosis (T score < -2.5 standard deviation). CONCLUSIONS: All aromatase inhibitor regimens reduced BMD. The sequential schedules were as detrimental for bone density as L monotherapy.

Prognostic and predictive value of centrally reviewed Ki-67 labeling index in postmenopausal women with endocrine-responsive breast cancer: results from Breast International Group Trial 1-98 comparing adjuvant tamoxifen with letrozole.

PURPOSE: To evaluate the prognostic and predictive value of Ki-67 labeling index (LI) in a trial comparing letrozole (Let) with tamoxifen (Tam) as adjuvant therapy in postmenopausal women with early breast cancer. PATIENTS AND METHODS: Breast International Group (BIG) trial 1-98 randomly assigned 8,010 patients to four treatment arms comparing Let and Tam with sequences of each agent. Of 4,922 patients randomly assigned to receive 5 years of monotherapy with either agent, 2,685 had primary tumor material available for central pathology assessment of Ki-67 LI by immunohistochemistry and had tumors confirmed to express estrogen receptors after central review. The prognostic and predictive value of centrally measured Ki-67 LI on disease-free survival (DFS) were assessed among these patients using proportional hazards modeling, with Ki-67 LI values dichotomized at the median value of 11%. RESULTS: Higher values of Ki-67 LI were associated with adverse prognostic factors and with worse DFS (hazard ratio [HR; high:low] = 1.8; 95% CI, 1.4 to 2.3). The magnitude of the treatment benefit for Let versus Tam was greater among patients with high tumor Ki-67 LI (HR [Let:Tam] = 0.53; 95% CI, 0.39 to 0.72) than among patients with low tumor Ki-67 LI (HR [Let:Tam] = 0.81; 95% CI, 0.57 to 1.15; interaction P = .09). CONCLUSION: Ki-67 LI is confirmed as a prognostic factor in this study. High Ki-67 LI levels may identify a patient group that particularly benefits from initial Let adjuvant therapy.

Comparing interval breast cancer rates in Norway and North Carolina: results and challenges.

OBJECTIVE: To compare interval breast cancer rates (ICR) between a biennial organized screening programme in Norway and annual opportunistic screening in North Carolina (NC) for different conceptualizations of interval cancer. SETTING: Two regions with different screening practices and performance. METHODS: 620,145 subsequent screens (1996-2002) performed in women aged 50-69 and 1280 interval cancers were analysed. Various definitions and quantification methods for interval cancers were compared. RESULTS: ICR for one year follow-up were lower in Norway compared with NC both when the rate was based on all screens (0.54 versus 1.29 per 1000 screens), negative final assessments (0.54 versus 1.29 per 1000 screens), and negative screening assessments (0.53 versus 1.28 per 1000 screens). The rate of ductal carcinoma in situ was significantly lower in Norway than in NC for cases diagnosed in both the first and second year after screening. The distributions of histopathological tumour size and lymph node involvement in invasive cases did not differ between the two regions for interval cancers diagnosed during the first year after screening. In contrast, in the second year after screening, tumour characteristics remained stable in Norway but became prognostically more favorable in NC. CONCLUSION: Even when applying a common set of definitions of interval cancer, the ICR was lower in Norway than in NC. Different definitions of interval cancer did not influence the ICR within Norway or NC. Organization of screening and screening performance might be major contributors to the differences in ICR between Norway and NC.

Robust vaccine-elicited cellular immune responses in breast milk following systemic Simian Immunodeficiency Virus DNA prime and live virus vector boost vaccination of lactating rhesus monkeys.

Breast milk transmission of HIV remains an important mode of infant HIV acquisition. Enhancement of mucosal HIV-specific immune responses in milk of HIV-infected mothers through vaccination may reduce milk virus load or protect against virus transmission in the infant gastrointestinal tract. However, the ability of HIV/SIV strategies to induce virus-specific immune responses in milk has not been studied. In this study, five uninfected, hormone-induced lactating, Mamu A*01(+) female rhesus monkey were systemically primed and boosted with rDNA and the attenuated poxvirus vector, NYVAC, containing the SIVmac239 gag-pol and envelope genes. The monkeys were boosted a second time with a recombinant Adenovirus serotype 5 vector containing matching immunogens. The vaccine-elicited immunodominant epitope-specific CD8(+) T lymphocyte response in milk was of similar or greater magnitude than that in blood and the vaginal tract but higher than that in the colon. Furthermore, the vaccine-elicited SIV Gag-specific CD4(+) and CD8(+) T lymphocyte polyfunctional cytokine responses were more robust in milk than in blood after each virus vector boost. Finally, SIV envelope-specific IgG responses were detected in milk of all monkeys after vaccination, whereas an SIV envelope-specific IgA response was only detected in one vaccinated monkey. Importantly, only limited and transient increases in the proportion of activated or CCR5-expressing CD4(+) T lymphocytes in milk occurred after vaccination. Therefore, systemic DNA prime and virus vector boost of lactating rhesus monkeys elicits potent virus-specific cellular and humoral immune responses in milk and may warrant further investigation as a strategy to impede breast milk transmission of HIV.

Global DNA hypomethylation coupled to repressive chromatin domain formation and gene silencing in breast cancer.

While genetic mutation is a hallmark of cancer, many cancers also acquire epigenetic alterations during tumorigenesis including aberrant DNA hypermethylation of tumor suppressors, as well as changes in chromatin modifications as caused by genetic mutations of the chromatin-modifying machinery. However, the extent of epigenetic alterations in cancer cells has not been fully characterized. Here, we describe complete methylome maps at single nucleotide resolution of a low-passage breast cancer cell line and primary human mammary epithelial cells. We find widespread DNA hypomethylation in the cancer cell, primarily at partially methylated domains (PMDs) in normal breast cells. Unexpectedly, genes within these regions are largely silenced in cancer cells. The loss of DNA methylation in these regions is accompanied by formation of repressive chromatin, with a significant fraction displaying allelic DNA methylation where one allele is DNA methylated while the other allele is occupied by histone modifications H3K9me3 or H3K27me3. Our results show a mutually exclusive relationship between DNA methylation and H3K9me3 or H3K27me3. These results suggest that global DNA hypomethylation in breast cancer is tightly linked to the formation of repressive chromatin domains and gene silencing, thus identifying a potential epigenetic pathway for gene regulation in cancer cells.

Bone marrow-derived cells are implicated as a source of lymphatic endothelial progenitors in human breast cancer.

Bone marrow-derived endothelial progenitor cells (EPCs) infiltrate into sites of neovascularization in adult tissues and mature into functional blood endothelial cells (BECs) during a process called vasculogenesis. Human marrow-derived EPCs have recently been reported to display a mixed myeloid and lymphatic endothelial cell (LEC) phenotype during inflammation-induced angiogenesis; however, their role in cancer remains poorly understood. We report the in vitro differentiation of human cord blood CD133(+)CD34(+) progenitors into podoplanin(+) cells expressing both myeloid markers (CD11b, CD14) and the canonical LEC markers vascular endothelium growth factor receptor 3 (VEGFR-3), lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1), and prospero homeobox 1 (PROX-1). These podoplanin(+) cells displayed sprouting behavior comparable to that of LECs in vitro and a dual hemangiogenic and lymphangiogenic activity in vivo in an endothelial cell sprouting assay and corneal vascularization assay, respectively. Furthermore, these cells expressed vascular endothelium growth factor (VEGF) family members A, -C, and -D. Thus, bone-marrow derived EPCs stimulate hemangiogenesis and lymphangiogenesis through their ability to differentiate into LECs and to produce angiogenic factors. Importantly, plasma from patients with breast cancer induced differentiation of CD34(+) cord blood progenitors into hemangiogenic and lymphangiogenic CD11b(+) myeloid cells, whereas plasma from healthy women did not have this effect. Consistent with these findings, circulating CD11b(+) cells from breast cancer patients, but not from healthy women, displayed a similar dual angiogenic activity. Taken together, our results show that marrow-derived EPCs become hemangiogenic and lymphangiogenic upon exposure to cancer plasma. These newly identified functions of bone-marrow derived EPCs are expected to influence the diagnosis and treatment of breast cancer.

The Oncogene Metadherin Modulates the Apoptotic Pathway Based on the Tumor Necrosis Factor Superfamily Member TRAIL (Tumor Necrosis Factor-related Apoptosis-inducing Ligand) in Breast Cancer.

Metadherin (MTDH), the newly discovered gene, is overexpressed in more than 40% of breast cancers. Recent studies have revealed that MTDH favors an oncogenic course and chemoresistance. With a number of breast cancer cell lines and breast tumor samples, we found that the relative expression of MTDH correlated with tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) sensitivity in breast cancer. In this study, we found that knockdown of endogenous MTDH cells sensitized the MDA-MB-231 cells to TRAIL-induced apoptosis both in vitro and in vivo. Conversely, stable overexpression of MTDH in MCF-7 cells enhanced cell survival with TRAIL treatment. Mechanically, MTDH down-regulated caspase-8, decreased caspase-8 recruitment into the TRAIL death-inducing signaling complex, decreased caspase-3 and poly(ADP-ribose) polymerase-2 processing, increased Bcl-2 expression, and stimulated TRAIL-induced Akt phosphorylation, without altering death receptor status. In MDA-MB-231 breast cancer cells, sensitization to TRAIL upon MTDH down-regulation was inhibited by the caspase inhibitor Z-VAD-fmk (benzyloxycarbonyl-VAD-fluoromethyl ketone), suggesting that MTDH depletion stimulates activation of caspases. In MCF-7 breast cancer cells, resistance to TRAIL upon MTDH overexpression was abrogated by depletion of Bcl-2, suggesting that MTDH-induced Bcl-2 expression contributes to TRAIL resistance. We further confirmed that MTDH may control Bcl-2 expression partly by suppressing miR-16. Collectively, our results point to a protective function of MTDH against TRAIL-induced death, whereby it inhibits the intrinsic apoptosis pathway through miR-16-mediated Bcl-2 up-regulation and the extrinsic apoptosis pathway through caspase-8 down-regulation.

Methylation signature of lymph node metastases in breast cancer patients.

BACKGROUND: Invasion and metastasis are two important hallmarks of malignant tumors caused by complex genetic and epigenetic alterations. The present study investigated the contribution of aberrant methylation profiles of cancer related genes, APC, BIN1, BMP6, BRCA1, CST6, ESR-b, GSTP1, P14 (ARF), P16 (CDKN2A), P21 (CDKN1A), PTEN, and TIMP3, in the matched axillary lymph node metastasis in comparison to the primary tumor tissue and the adjacent normal tissue from the same breast cancer patients to identify the potential of candidate genes methylation as metastatic markers. METHODS: The quantitative methylation analysis was performed using the SEQUENOM's EpiTYPER? assay which relies on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). RESULTS: The quantitative DNA methylation analysis of the candidate genes showed higher methylation proportion in the primary tumor tissue than that of the matched normal tissue and the differences were significant for the APC, BIN1, BMP6, BRCA1, CST6, ESR-b, P16, PTEN and TIMP3 promoter regions (P<0.05). Among those candidate methylated genes, APC, BMP6, BRCA1 and P16 displayed higher methylation proportion in the matched lymph node metastasis than that found in the normal tissue (P<0.05). The pathway analysis revealed that BMP6, BRCA1 and P16 have a role in prevention of neoplasm metastasis. CONCLUSIONS: The results of the present study showed methylation heterogeneity between primary tumors and metastatic lesion. The contribution of aberrant methylation alterations of BMP6, BRCA1 and P16 genes in lymph node metastasis might provide a further clue to establish useful biomarkers for screening metastasis.