Breast carcinomas are often infiltrated by inflammatory cells, particularly macrophages and T lymphocytes, but the significance of these cells remains unclear. One possible role of these inflammatory cells is that they represent a cell-mediated immune response against the carcinoma. CD8(+) lymphocytes are a known crucial component of cell-mediated immunity. The purpose of this study was to explore the prognostic value of tumor-infiltrating CD8(+) cytotoxic lymphocytes in breast cancer. Tumor-infiltrating CD8(+) lymphocytes were assessed by immunohistochemical staining of tissue microarray cores from 1,334 unselected breast tumors from patients with long-term follow-up. The number of CD8(+) T cells was counted in tumor nests (intratumoral), in stroma adjacent to tumor cells, and in stroma distant to tumor cells, and their relationship with clinical outcome was determined. The total number of CD8(+) cells was positively correlated with tumor grade (r(s) = 0.20; P < .001) and inversely correlated with patient's age at diagnosis, estrogen receptor-alpha (ER-α), and progesterone receptor (PgR) expression (Mann-Whitney U test, P < .001). The total patient cohort was randomly divided into two separate training and validation sets before performing univariate survival analysis. Total number and distant stromal CD8(+) lymphocytes were associated with better patient survival (P = .041 and P < .001, respectively) in the training set. In multivariate analysis, total CD8(+) T-cell count was an independent prognostic factor in both training and validation sets. These results suggest that tumor-infiltrating CD8(+) T lymphocytes have antitumor activity as judged by their favorable effect on patients' survival and could potentially be exploited in the treatment of breast cancer.

Mucins are a large family of glycoproteins expressed by many epithelial cells and their malignant counterparts. Much interest has been focused on expression of its members in breast cancer because of their potential role as prognostic indicators and their involvement in cancer therapy. We have examined 1447 cases of invasive breast carcinoma with a long-term follow-up, using tissue microarray (TMA) technology and immunohistochemistry to evaluate the expression profiles of several mucins (MUC1, MUC2, MUC3, MUC4, MUC5AC and MUC6) and to assess their prognostic value. We detected MUC1 expression in 91% of tumours. MUC1 overexpression was associated with a lower grade, smaller tumour size, a higher oestrogen receptor (ER)-positive phenotype and absence of both regional recurrence and distance metastasis. The subcellular localization but not the level of expression had a prognostic value in predicting outcome. The aberrant cytoplasmic and membranous localization of MUC1 was associated with poor outcome compared with apical localization, which is the normal physiological site of expression. MUC2 expression was noticed in only 8.3% of all cases and was restricted to the cytoplasm of the tumour cells. An inverse trend was identified between MUC2 expression and lymph node stage and vascular invasion status. On excluding cases of mucinous carcinoma from the analysis, the inverse association with vascular invasion was still defined and in addition an inverse association with ER status emerged. MUC3 expression was detected in 91% of cases and its expression was associated with increased local recurrence, and lymph node stage. The membranous expression of MUC3 was found to be a potentially poor prognostic feature, with higher grade and poorer Nottingham Prognostic Index (NPI), and negative ER expression. MUC4, MUC5AC and MUC6 were expressed in 95, 37 and 20% of cases, respectively. Apart from an association between MUC4 expression and tumour grade and between MUC6 and ER-negative tumours, no other associations with any clinicopathological variables were found. Apart from the higher expression of MUC2 and MUC6 in mucinous carcinomas, no association was found between the expression of different mucins and tumour type. No association between the level of expression of any of the studied mucins and patient outcomes has been identified. In conclusion, most breast carcinomas express MUC1, MUC3 and MUC4. Among the various mucins expressed in breast cancer, MUC1 and MUC3 are potential prognostic indicators, MUC1 having the strongest relationship with patient outcome.

ER+/PgR-and ER-/PgR+ tumors are biologically and clinically distinct groups of breast cancer that may require different treatment strategies with ER-/PgR+ exhibiting more aggressive behavioral characteristics.

Macrophages are heterogeneous with different subsets having different functions. The present study suggests that overall macrophage numbers are not related to prognosis in breast cancer. However, further studies are needed to investigate the potential role of different subsets of macrophages.

Although the favourable role of T lymphocyte populations in different tumour types is established, that of B cells is still a matter of debate and needs further clarification. The presence of tumour-infiltrating B cells may represent an antibody response against breast tumour antigens. We used immunohistochemistry to investigate the density and localisation of B lymphocytes infiltrating 1470 breast tumours and to identify any prognostic significance and relationship to various clinicopathological factors. Higher numbers of CD20(+) cells were found in the stroma away from the carcinoma (mean 12 cells) compared with either intratumoural or adjacent stromal compartments (mean 1 cell). The majority of tumours showed a diffuse pattern of B cells rather than aggregates. There was a positive correlation between higher numbers of total CD20(+) B cells and higher tumour grade (r (s) = 0.20, P < 0.001), ER and PgR negativity (P < 0.001), and basal phenotype (P < 0.001) subclass. In univariate survival analysis, higher total number of infiltrating CD20(+) cells, irrespective of location, was associated with significantly better BCSS (P = 0.037) and longer DFI (P = 0.001). In multivariate analysis, total CD20(+) B cell count (HR = 0.75, 95% CI = 0.58-0.96 for BCSS and HR = 0.72, 95% CI = 0.58-0.89, for DFI), tumour size, nodal stage, grade, vascular invasion, HER-2 status, and total CD8(+) T cell count were independently associated with outcome. This suggests that humoral immunity, in addition to the cell mediated immunity, may be important in breast cancer. This should be considered in breast cancer immunotherapy and vaccine strategies.

Introduction IL-17 plays an important role in autoimmunity, promoting autoimmunity, inflammation and invasion in multiple sclerosis, rheumatoid arthritis and type I diabetes. The role of IL-17 in cancer is unclear, however, as there are few studies examining IL-17 protein expression in cancer. We therefore examined IL-17 protein expression in human breast cancer and modelled its potential biological significance in vitro.

The presence of vascular invasion (VI), encompassing both lymphovascular invasion (LVI) and blood vascular invasion (BVI), in breast cancer has been found to be a poor prognostic factor. It is not clear, however, which type of VI plays the major role in metastasis. The aims of this study were to use an endothelial subtype specific immunohistochemical approach to distinguish between LVI and BVI by comparing the differential expression of blood vascular (CD34 and CD31) and lymphatic markers (podoplanin/D2-40) to determine their prognostic role in a well-characterized group of breast cancer patients with known long-term follow-up. Sections from 177 consecutive paraffin-embedded archival specimens of primary invasive breast cancer were stained for expression of podoplanin, D2-40, CD31, and CD34. BVI and LVI were identified and results were correlated with clinicopathologic criteria and patient survival. VI was detected in 56/177 specimens (31.6%); 54 (96.4%) were LVI and 2 (3.5%) were BVI. The presence of LVI was significantly associated with the presence of lymph node metastasis, larger tumor size, development of distant metastasis, regional recurrence and worse disease-free interval and overall survival. In multivariate analysis, LVI retained significance association with decreased disease-free interval and overall survival. In conclusion, VI in breast cancer is predominantly of lymph vessels and is a powerful independent prognostic factor, which is associated with risk of recurrence and death from the disease. The use of immunohistochemical staining with a lymphendothelial specific marker such as podoplanin/D2-40 increases the accuracy of identification of patients with tumor associated LVI.

Vascular endothelial cell growth factors (VEGF)-A, -C and -D have potent angio and lymphangiogenic functions in experimental models, although their role in the progression of human breast cancer is unclear. The aims of the current study were to examine the relationship between the expression of the aforementioned growth factors with the angio and lymphangiogenic characteristics of breast cancer, and to assess their suitability as potential prognostic factors. Paraffin-embedded sections of 177 primary invasive breast cancer, with complete clinical follow-up information for 10 years, were stained for VEGF-A, -C, -D, podoplanin and CD34 using standard immunohistochemical approaches. The expression of the growth factors was correlated with clinicopathological criteria and patients' survival. Lymph vessel density (LVD) and microvessel density (MVD) were assessed and correlated with expression of the growth factors. Vascular endothelial cell growth factor-A, -C and -D were highly expressed in 40, 37 and 42% of specimens, respectively. High expression of VEGF-A and -C, but not of -D, was associated with a higher LVD (P ¼ 0.013 and P ¼ 0.014, respectively), a higher MVD (Po0.001 and P ¼ 0.002, respectively), the presence of lymph node metastasis (Po0.001 and Po0.001, respectively), distant metastasis (P ¼ 0.010 and P ¼ 0.008, respectively) and a shorter Overall Survival (P ¼ 0.029 and 0.028, respectively). In conclusion, breast cancers that express high levels of VEGF-A and -C are characterised by a poor prognosis, likely through the induction of angio and lymphangiogenesis. Examination of expression of VEGF-A and -C in breast cancer may be beneficial in the identification of a subset of tumours that have a higher probability of recurrence and metastatic spread.