Neutrophil-lymphocyte ratio reflects tumour-infiltrating lymphocytes and tumour-associated macrophages and independently predicts poor outcome in breast cancers with neoadjuvant chemotherapy.

AIMS: The neutrophil-lymphocyte ratio (NLR) is a systemic reflection of cancer-associated inflammation and a prognostic marker for breast cancer. For the local tumour microenvironment, tumour-infiltrating lymphocytes (TILs) and tumour-associated macrophages (TAMs) are also highly correlated with breast cancer survival. This study aimed to explore the relationship between the circulating and local immune microenvironment, and to further delineate the prognostic role of NLR in breast cancer patients receiving neoadjuvant chemotherapy (NAC). METHODS: A cohort of breast cancer patients receiving NAC with subsequent surgery was retrieved. Clinical data were reviewed. Histological slides and CD8 immunohistochemistry from biopsy (pre-chemotherapy) and excision (postchemotherapy) specimens were assessed for TILs and TAMs. RESULTS: A total of 146 patients were included. There was a significant positive correlation between pre- and postsurgery NLR at a cut-off of 2.6 (median pre-chemotherapy NLR) (P < 0.001). NLR pre-chemotherapy was associated positively with necrosis on biopsy (P = 0.027) and excision (P = 0.021) and TAMs on excision (P = 0.049). NLR 1 year postsurgery was associated with high tumour stage (P = 0.050) and low histological grade (P = 0.008). TIL count was lower in NLR-high cases at almost all time-points by histological assessment and CD8 immunostaining (P < 0.050). In multivariate analysis, postsurgery NLR is an independent predictor for overall survival [OS; hazard ratio (HR) = 9.524, P < 0.001], breast cancer-specific survival (BCSS) (HR = 10.059, P = 0.001) and disease-free survival (DFS; HR = 2.824, P = 0.016). CONCLUSIONS: The association between NLR with tumour necrosis, TAMs and TILs illustrates an interaction between the circulating and local immune microenvironment. Late NLR is a strong indicator of outcome and may be useful for prognostication and disease monitoring.

SALL4 promotes tumor progression in breast cancer by targeting EMT.

Sal-like protein 4 (SALL4) is overexpressed in breast cancer and might contribute to breast cancer progression, but the molecular mechanism remains unknown. Here, we found that within a group of 371 ethnic Chinese breast cancer patients, SALL4 was associated with lower grade (P = .002) and progesterone receptor positivity (P = .004) for overall cases; lower Ki67 (P = .045) and high vimentin (P = .007) for luminal cases. Patients with high SALL4 expression in lymph node metastasis showed a significantly worse survival than those with low expression. Knockout of SALL4 in a triple-negative breast cancer cell line MDA-MB-231-Red-FLuc-GFP led to suppressed ability in proliferation, clonogenic formation, migration, and mammosphere formation in vitro, tumorigenicity and lung colonization in vivo. On the other hand, overexpression of SALL4 enhanced migration and mammosphere formation in vitro and tumorigenicity in vivo. Mechanistically, there was a positive correlation between SALL4 expression and mesenchymal markers including Zinc finger E-box binding homeobox 1 (ZEB1), vimentin, Slug, and Snail in vivo. Chromatin immunoprecipitation experiment indicated that SALL4 can bind to the promoter region of vimentin (-778 to -550 bp). Taken together, we hypothesize that SALL4 promotes tumor progression in breast cancer by inducing the mesenchymal markers like vimentin through directly binding to its promoter. Increased SALL4 level in metastatic lymph node relative to the primary site is an important poor survival marker in breast cancer.

Molecular Classification of Breast Cancer.

Cancer classification aims to provide an accurate diagnosis of the disease and prediction of tumor behavior to facilitate oncologic decision making. Traditional breast cancer classification, mainly based on clinicopathologic features and assessment of routine biomarkers, may not capture the varied clinical courses of individual breast cancers. The underlying biology in cancer development and progression is complicated. Recent findings from high-throughput technologies added important information with regard to the underlying genetic alterations and the biological events in breast cancer. The information provides insights into new treatment strategies and patient stratifications that impact on the management of breast cancer patients. This review provides an overview of recent data on high throughput analysis of breast cancers, and it analyzes the relationship of these findings with traditional breast cancer classification and their clinical potentials.

Expression of biomarkers in the AKT pathway correlates with malignancy and recurrence in phyllodes tumours of the breast.

AIMS: Phyllodes tumours (PTs) of the breast are uncommon fibroepithelial neoplasms with the potential to recur and metastasise. Apart from histological grading, the expression of biological markers and its relationship with tumour behaviour have been topics of interest. The phosphatidylinositol 3-kinase (PI3K)-AKT pathway regulates diverse biological functions, and is one of the most frequently deregulated pathways in cancers. Little is known of PI3K-AKT pathway alteration in PT. We aim to investigate the alterations in different component of AKT pathway in PTs. METHODS AND RESULTS: This study investigated the expression of four biological markers involved in this pathway (PTEN, INPP4B, PI3KCA and pAKT) in 134 PTs by the use of immunohistochemistry. According to an immunoscore incorporating staining intensity and proportion, low epithelial INPP4B expression (P = 0.045) was associated with recurrence. A trend of association was found for low epithelial PTEN expression with recurrence (P = 0.090). Interestingly, low epithelial INPP4B expression was also associated recurred tumours (P = 0.043). Stromal PI3KCA expression (P = 0.016) and pAKT expression (P = 0.006) were found to be correlated with increased histological grade, but an opposite trend was seen for stromal INPP4B expression (P = 0.018). In addition, epithelial and stromal PTEN expression, PI3KCA expression and pAKT expression showed strong correlations with each other (P ≤ 0.001). CONCLUSIONS: These findings indicate that alterations in AKT pathway activation may correlate with malignant transformation and recurrence in PT. Low epithelial INPP4B/PTEN expression is associated with shorter recurrence-free survival. These observations suggest that the pathway may play a crucial role in the biological behaviour and progression of PT, and assessing the expression of this pathway may be of value in diagnosis, grading, prognostication, and management.

Diagnostic upgrade of atypical ductal hyperplasia of the breast based on evaluation of histopathological features and calcification on core needle biopsy.

AIMS: Atypical ductal hyperplasia (ADH) of breast is increasingly diagnosed in core needle biopsy (CNB). As higher-grade lesions were found in the excision in a substantial proportion of ADH on CNB, factors predicting risk of subsequent upgrade are clinically significant. This study aims to investigate relevant histopathological factors in CNB that could predict diagnostic upgrade at excision. METHODS AND RESULTS: One hundred and forty-three cases of CNB with paired subsequent excision were evaluated for multiple clinicopathological parameters related to CNB sampling, ADH morphology, calcification and other co-existing histological features, and which of these parameters were associated with diagnostic upgrade at subsequent excisions were determined. Forty-eight cases (34.3%) were upgraded to malignancy, including 15 invasive cancers and 33 ductal carcinomas in situ (DCIS). An increased tissue area occupied by ADH (P = 0.026), a higher number of ADH foci (P = 0.004), the presence of solid pattern (P = 0.037) and older age (P = 0.012) were positively associated with upgrade, while negative associations were found with the presence of micropapillary pattern (P = 0.025), co-existing columnar cell lesions (CCL) (P = 0.001) and the presence of calcifications (P = 0.009). Multivariate logistic regression analysis showed that the number of ADH foci (HR = 2.810, P = 0.013) was an independent positive predictor, while co-existing CCL (HR = 0.391, P = 0.013) was an independent negative predictor for upgrade. CONCLUSIONS: Patients with ADH in CNB showing the presence of co-existing CCL and a lower number of ADH foci have a lower risk of disease upgrade at excision, and are potential candidates for observation-only management.

Amyloid Precursor Protein Is Associated with Aggressive Behavior in Nonluminal Breast Cancers.

BACKGROUND: ß-amyloid precursor protein (APP), a potential target for Alzheimer's disease treatment, has recently been shown to take part in carcinogenesis. Increased APP promotes migration, survival, and proliferation in breast cancer cell lines. We examined the clinical value of APP in breast cancers. A comprehensive examination of clinicopathological features related to APP expression in a large cohort of breast cancers and the corresponding metastatic lymph nodes was performed. APP expression and its prognostic impact in different breast cancer subtypes were examined. RESULTS: APP was highly expressed in nonluminal breast cancers and correlated with features associated with nonluminal breast cancers (including higher grade, the presence of necrosis, and higher proliferative index, growth factor receptor, and basal marker expression). Multivariate Cox hazard analysis demonstrated that APP was an independent adverse prognostic factor of disease-free survival (DFS; hazard ratio [HR], 2.090; p = .013; 95% confidence interval [CI], 1.165-3.748) and breast cancer-specific survival (BCSS; HR, 2.631; p = .002; 95% CI, 1.408-4.915) in the nonluminal group. The independent prognostic impact was also seen in triple negative breast cancers. Interestingly, a higher expression of APP was found in nodal metastasis compared with primary tumor. Such APP upregulation was correlated with further distal metastasis and poorer outcome (DFS: log-rank, 12.848; p < .001; BCSS: log-rank, 13.947; p < .001). CONCLUSION: Our findings provided evidence of oncogenic roles of APP in clinical breast cancers. Patients with positive APP expression, particularly those with APP upregulation in lymph node metastases, may require vigilant monitoring of their disease and more aggressive therapy. IMPLICATIONS FOR PRACTICE: ß-amyloid precursor protein (APP), a potential target for Alzheimer's disease, has recently been implicated in oncogenesis. Here, evidence of its roles in clinical breast cancers is provided. Positive APP expression was found to be an independent prognostic factor in nonluminal cancers, particularly triple negative breast cancers (TNBCs). Interestingly, a higher APP in nodal metastases was associated with distal metastases. TNBCs are heterogeneous and currently have no available target therapy. APP could have therapeutic potential and be used to define the more aggressive cases in TNBCs. Current prognostic analysis is based on primary tumor. The present data suggest that investigation of nodal metastases could provide additional prognostic value.

GATA-3 is superior to GCDFP-15 and mammaglobin to identify primary and metastatic breast cancer.

PURPOSE: Despite numerous studies on the utility of GATA-3 as breast cancer marker, its comparison with other breast markers, its concordance between primary and metastatic tumors and its expression in primary cancers from sites with frequent breast metastases remains unclear. METHODS: To address these questions, totally 993 invasive breast cancers (IBC), 254 paired nodal metastases, 23 distant metastases, and 208 lung carcinomas were included. GATA-3 expression was analyzed by immunohistochemistry and compared to other breast markers [gross cystic disease fluid protein 15 (GCDFP-15) and mammaglobin (MGB)]. RESULTS: GATA-3 was expressed in 82.5% of IBC, predominantly in luminal (93.9%), and lower in non-luminal cancers [59.6% of HER2 overexpressing (HER2-OE) and 38.1% of triple negative breast cancer (TNBC) subtypes]. GATA-3 identified more IBC than GCDFP-15 (23.9%) and MGB (46.6%). However, MGB showed a comparable sensitivity for non-luminal cancers to GATA-3. Combining MGB and GATA-3 improved sensitivity for both HER2-OE (80.8%) and TNBC cases (55.4%). GATA-3 showed a high sensitivity for nodal metastases and distant metastases, with good concordance with primary tumors. GATA-3 was expressed in 1.0% of lung carcinomas, with sensitivity and specificity of 82.5 and 99.0% in differentiating IBC and lung carcinoma. CONCLUSIONS: GATA-3 expression was the highest in luminal breast carcinomas, and showed higher sensitivity than GCDFP-15 and MGB. However, in the poorly differentiated IBC, its utility was still limited. One should be aware of the possible GATA-3 expression in lung carcinomas.

Intratumoral Heterogeneity in Breast Cancer: A Comparison of Primary and Metastatic Breast Cancers.

Intratumoral heterogeneity presents challenges in the management of cancer. To gain deeper insight in intratumoral heterogeneity at different levels and tumor sites for common biomarkers in breast cancers, this report examines seven cases of invasive breast cancer with multiple axillary nodal metastases and/or recurrences for immunohistochemical expression of estrogen receptors, progesterone receptors, human epidermal growth receptor 2, and Ki67 on all tissue blocks in both primary and metastatic tumors. The Oncologist 2017;22:487-490.

Distinct Tertiary Lymphoid Structure Associations and Their Prognostic Relevance in HER2 Positive and Negative Breast Cancers.

BACKGROUND: The presence of tumor infiltrating lymphocytes (TIL) is associated with favorable prognosis. Recent evidence suggested that not only their density, but also the spatial organization as tertiary lymphoid structures (TLS), play a key role in determining patient survival. MATERIALS AND METHODS: In a cohort of 248 breast cancers, the clinicopathologic association and prognostic role of TLS was examined. RESULTS: Tertiary lymphoid structures were associated with higher tumor grade, apocrine phenotype, necrosis, extensive in situ component, lymphovascular invasion (LVI), and high TIL. For biomarkers, TLS were associated with hormone receptors negativity, HER2 positivity, and c-kit expression. Tertiary lymphoid structures were significantly related to better disease-free survival (DFS) in HER2 positive (HER2+) breast cancers (log-rank = 4.054), which was not dependent on high TIL status. The combined TLS and TIL status was an independent favorable factor associated with DFS in those cases. Interestingly, tumor cell infiltration into the TLS was found in 41.9% of TLS positive cases. It was associated with LVI in HER2 negative (HER2-) TLS positive (particularly estrogen receptor positive [ER+] HER2-) cases. In the ER+ HER2- cases, tumor cell infiltration into TLS was also associated with increased pathologic nodal stage (pN) stage and nodal involvement. CONCLUSION: Tertiary lymphoid structures showed a similar relationship with clinicopathologic features and biomarkers as TIL. The presence of TLS, irrespective of TIL level, could be an important favorable prognostic indicator in HER2+ breast cancer patients. Given the significance of TLS in promoting effective antitumor immunity, further understanding of its organization and induction may provide new opportunities to improve the current immunotherapy strategies. IMPLICATIONS FOR PRACTICE: Despite recent interest on the clinical value of tumor infiltrating lymphocyte (TIL), little was known on the clinical significance on their spatial organization as tertiary lymphoid structures (TLS). Although TLS showed similar relationships with clinicopathologic features and biomarkers as TIL, the prognostic value of TLS, particularly in HER2 positive cancers, was independent of TIL. Moreover, tumor infiltration could be present in TLS which appears to be related to tumor invasion in HER2 negative cancers. Overall, the results demonstrated the additional value for TLS in HER2 cancer subtypes. Further investigations and its standardized evaluation will enhance its use as standard practice.

PD-L1 expression and tumor infiltrating PD-1+ lymphocytes associated with outcome in HER2+ breast cancer patients.

PURPOSE: Clinical trials showing programmed death (PD)-1-PD-ligand 1 (L1) axis as a promising therapeutic target in melanoma and non-small cell lung cancers have made the pathway a focus of recent attention. However, the data regarding PD-L1/PD-1 in breast cancer are inconsistent. Given the heterogeneity of breast cancers, the clinical relevance of PD-L1 and PD-1 tumor infiltrating lymphocytes (TIL) may vary according to subtypes of breast cancer. We aim to investigate PD-L1 expression in a large cohort of breast cancers and analyze its clinico-pathological as well as outcome relationship according to molecular subtypes. Also, we evaluate the relationship of PD-1 TIL and PD-L1 expression with patients' survival, particularly for breast cancers with high TIL. METHODS AND RESULTS: Immunohistochemical analysis of PD-L1 on tissue arrays for 1091 breast cancer patients and PD-1 TIL on 97 whole sections was performed. Associations of PD-L1 with luminal cancers (p < 0.001) and features associated with that subtype [lower histologic grade, absence of necrosis, ER, PR, and AR expression (p < 0.001)] were observed. However, in HER2+ breast cancers, PD-L1 was an independent poor prognostic indicator (DFS: HR = 1.866, p = 0.001; OS: HR = 1.517, p = 0.036). Interestingly, HER2+ cancers showed a lower PD-1 TIL level compared to the other high TIL cases (p = 0.011). Cases with low PD-TIL but high PD-L1 expression showed the worst survival. This could be indicative of an active immune suppression by PD-L1 expression. CONCLUSIONS: Our data showed the relevance of PD-L1 expression in HER2+ breast cancer. A combined evaluation of PD-L1 and PD-1 TIL in the prognosis of breast cancer might also be of value in treatment prediction.

Expression and Clinical Significance of Herpes Virus Entry Mediator (HVEM) in Breast Cancer.

BACKGROUND: Immune checkpoint blockades are currently actively investigated in invasive breast cancers. Given the complexity of immune regulation, multiple inhibitory molecules within the immune checkpoint framework would be involved in tumor immune escape. Evaluation of the components within the framework is a prerequisite for not only identification of additional treatment targets and optimization of immunotherapeutic strategies but also understanding the prognostic value of these molecules. METHODS AND RESULTS: We examined a recently described component, herpes virus entry mediator (HVEM), in a large cohort of invasive breast cancers using immunohistochemistry, and evaluated its clinical relevance. HVEM expression was associated with aggressive tumor features, namely high grade (p < 0.001), high pT (p = 0.001) and pN stage (p = 0.008), and was most prevalently found in human epidermal growth factor receptor 2-overexpressed subtype (67%). Interestingly, a negative association with programmed death-ligand 1 (p = 0.021) has been observed. The prognostic impact of HVEM depended on the level of tumor-infiltrating lymphocytes (TILs), with the worst outcome occurring in patients with low TIL, HVEM-positive tumors. CONCLUSION: HVEM plays significant oncogenic roles in breast carcinogenesis, and may also be a tumor-specific target.

Bcl2 and Ki67 refine prognostication in luminal breast cancers.

Combined B-cell lymphoma 2 (Bcl2) and Ki67 expression for breast cancer prognostication has been proposed recently. However, the combinatorial relationship with patient outcome, clinico-pathologic features, and various biomarkers has not been fully explored. Bcl2 and Ki67 expression were examined in a large cohort of breast cancers. Differential Bcl2 and Ki67 combinatorial analysis, particularly in luminal cancers, were evaluated with respect to the clinico-pathologic features, biomarkers profile and outcome. Combined Bcl2/Ki67 phenotypes classified by Bcl2 and Ki67 cutoffs showed a better correlation with outcome. Multivariate analysis revealed this to be an independent prognostic factor in luminal cancers. Both Ki67 and Bcl2 contributed to the prognostic implications of different subgroups defined by Bcl2/Ki67 combination phenotypes with clinico-pathologic features and biomarkers profile. Ki67low/Bcl2high cases showed better DFS (HR = 2.17, P = 0.015) and OS (HR = 3.217, P = 0.015) compared to Ki67high/Bcl2low cases. Interestingly, Ki67low/Bcl2high cases also showed better outcome than other phenotypes in grade 2 cancers (log-rank = 4.844, P = 0.028) and TNM stage 2 cancers (log-rank = 8.161, P = 0.004). This classification by Bcl2/Ki67 combination phenotypes, together with PR expression, can also refine luminal A cancers prognostication. Not all PR low luminal A cases had poorer outcome compared to the PR high luminal A cases; poor prognosis was only limited to those with also low Bcl2 (log-rank = 23.568, P < 0.001 compared to PR high Bcl2 high cases). The combined Ki67/Bcl2 phenotyping was useful in luminal cancers prognostication. It also refined prognostication in intermediate groups (grade 2 and stage 2 cancers) of luminal cancers; and aided in further classification of luminal A cancers.

Anterior Gradient 2 is a Poor Outcome Indicator in Luminal Breast Cancer.

BACKGROUND: Recent data have shown anterior gradient 2 (AGR2) to be a novel oncogene overexpressed in luminal breast cancers. However, many studies only focused on its relationship with treatment resistance, and the clinical relevance of AGR2 has not been widely evaluated. METHODS: AGR2 expression in a cohort of breast cancer was correlated with the clinicopathologic features, biomarker expression, and patient survival. RESULTS: AGR2 expression correlated with lower grade (p = 0.002) and absence of necrosis (p = 0.001) and was most highly expressed in luminal cancers (p < 0.001). Correspondingly, AGR2 correlated positively with estrogen receptor, progesterone receptor, and androgen receptor (p < 0.001) and negatively with epidermal growth factor receptor (p =0.002) and CK5/6 (p < 0.001). AGR2 was an independent unfavorable disease-free survival prognostic factor in patients with hormone therapy (hazard ratio 2.537, p = 0.023). Interestingly, it was also an independent adverse disease-free survival prognostic factor in node-positive luminal cancers (hazard ratio 3.381, p = 0.016). AGR2 expression was higher in nodal metastasis than the corresponding primary tumors in luminal cancers (p= 0 .023). CONCLUSIONS: The prognostic impact of AGR2 expression could be related to treatment outcome in estrogen receptor-positive breast cancers and/or its metastasis-promoting effects. It could be an important tumor biomarker and negative prognostic factor potentially exploitable in clinical practice.

GATA-binding protein 3, gross cystic disease fluid protein-15 and mammaglobin have distinct prognostic implications in different invasive breast carcinoma subgroups.

AIMS: To evaluate the prognostic significance of GATA-binding protein 3 (GATA-3), gross cystic disease fluid protein-15 (GCDFP-15) and mammaglobin (MGB) in invasive breast carcinomas (IBCs). METHODS AND RESULTS: GATA-3, GCDFP-15 and MGB were expressed in 37.9% (370/976), 26.0% (254/978) and 35.3% (348/986) of this cohort of 1017 IBCs, respectively. GCDFP-15 was an independent favourable prognostic factor in all cases [disease-free survival (DFS), hazard ratio (HR) 0.587, P = 0.049; overall survival (OS), HR 0.512, P = 0.049], as well as in oestrogen receptor (ER)-negative (DFS, HR 0.353, P = 0.012; OS, HR 0.310, P = 0.017) and HER2-positive (DFS, HR 0.279, P = 0.036; OS, HR 0.235, P = 0.050) cases; it also refined the prognostication of molecular apocrine cancers. GATA-3 and MGB did not show any prognostic significance. CONCLUSIONS: The commonly used breast carcinoma biomarkers vary in their prognostic implications. GCDFP-15 independently indicated a favourable prognosis, especially in ER-negative, HER2-positive and molecular apocrine cancers. GATA-3 and MGB were not associated with outcome.

MicroRNAs are differentially deregulated in mammary malignant phyllodes tumour.

AIMS: MicroRNAs (miRs) have been shown to play important roles in tumour progression. Their expression pattern can be useful for cancer classification. However, little is known about miRs in mammary phyllodes tumours (PT). METHODS AND RESULTS: In this study, polymerase chain reaction (PCR)-based miR profiling was performed in a small PT cohort to identify deregulated miRs in malignant PT. The purported roles and targets of these miRs were further validated. Unsupervised clustering of miR expression profiling segregated PT into different grades, implicating the miR profile in PT classification. Among the deregulated miRs, miR-21, miR-335 and miR-155 were validated to be higher in malignant than in lower-grade PT in the independent cohort by quantitative PCR (qPCR) (P ≤ 0.032). Their expression correlated with some of the malignant histological features, including high stromal cellularity, nuclear pleomorphism and mitosis. Subsequent analysis of their downstream proteins, namely PTEN for miR-21/miR-155 and Rb for miR-335, also showed an independent significant negative association between miR and protein expression. CONCLUSIONS: Differential expression of miRs in PT could be useful in diagnosis and grading of PT. Their deregulated expression, together with the altered downstream targets, implicated their active involvement in PT malignant transformation.

CD73 expression on extracellular vesicles derived from CD4+ CD25+ Foxp3+ T cells contributes to their regulatory function.

CD4(+)CD25(+)Foxp3(+) Treg cells maintain immunological tolerance. In this study, the possibility that Treg cells control immune responses via the production of secreted membrane vesicles, such as exosomes, was investigated. Exosomes are released by many cell types, including T cells, and have regulatory functions. Indeed, TCR activation of both freshly isolated Treg cells and an antigen-specific Treg-cell line resulted in the production of exosomes as defined morphologically by EM and by the presence of tetraspanin molecules LAMP-1/CD63 and CD81. Expression of the ecto-5-nucleotide enzyme CD73 by Treg cells has been shown to contribute to their suppressive function by converting extracellular adenosine-5-monophosphate to adenosine, which, following interaction with adenosine receptors expressed on target cells, leads to immune modulation. CD73 was evident on Treg cell derived exosomes, accordingly when these exosomes were incubated in the presence of adenosine-5-monophosphate production of adenosine was observed. Most importantly, CD73 present on Treg cell derived exosomes was essential for their suppressive function hitherto exosomes derived from a CD73-negative CD4(+) T-cell line did not have such capabilities. Overall our findings demonstrate that CD73-expressing exosomes produced by Treg cells following activation contribute to their suppressive activity through the production of adenosine.

Lymphocytic infiltrate is associated with favorable biomarkers profile in HER2-overexpressing breast cancers and adverse biomarker profile in ER-positive breast cancers.

The value for lymphocytic infiltration (LI) has been increasingly recognized for tumor assessment. In breast cancer, however, the overall significance of LI remains poorly defined, probably due to its heterogeneity. A large cohort of breast cancer was evaluated for the degree of LI and its association with traditional pathologic factors, biomarker expression, and cancer subtypes. The number of CD8 cytotoxic effector and FoxP3 regulatory T cell (Treg) was evaluated in those cases with high LI. High LI was associated with negative ER and PR but positive HER2 and EGFR expression (p < 0.001 for all). In ER-positive cancers, high LI was associated with poor prognostic features including higher grade, the presence of necrosis, and lymphovascular invasion (LVI) (p = 0.007 for LVI and <0.001 for the others). Conversely, LI correlated with smaller tumor size, a good prognostic feature (p = 0.046) in HER2+ ER-cancers. These observations suggested LI may show opposite prognostic values in different breast cancer subgroups. Interestingly, when the phenotype of LI in these subgroups was evaluated, a strong positive association with intratumoral accumulation of Treg was found in ER-positive cancers (p = 0.003, Rs = 0.319), while the opposite was observed in HER2+ ER-cancers (p < 0.001, Rs = -0.427). Also, in ER-positive cancers, positive associations between peri- and intra-tumoral distribution were found with both CD8 and Tregs (CD8: p < 0.001, Rs = 0.547; Treg: p = 0.001, Rs = 0.460). Nonetheless, in HER2+ ER-cancers, such strong association was found with CD8 (p < 0.001, Rs = 0.766) but not Tregs. The results may implicate a differential intratumoral migration of LI in different subtypes of breast cancer. In summary, the clinical value of LI in breast cancers could be subtype-dependent. In ER-positive cancers, high LI correlated with biologic parameters associated with poor prognosis, whereas in HER2 positive cancers, LI correlated with biologic parameters of favorable prognosis.

A novel morphologic-molecular recurrence predictive model refines traditional prognostic tools for invasive breast carcinoma.

BACKGROUND: Histologic grade, TNM stage, and Nottingham Prognostic Index are traditional prognostic tools for breast cancer. "IHC-molecular" classification of breast cancer can also identify patients at different recurrence risks and provides insight into cancer therapy. However, cancers in each group are heterogeneous. A model based on the comprehensive analysis of morphologic features and molecular subtype was constructed to predict recurrence and refine these traditional prognostic tools. METHODS: Morphologic features including histologic grade, fibrotic focus, extensive intraductal component, lymphocytic infiltrate, lymphovascular invasion, tumor necrosis, tumor margin and TNM stage, and molecular subtypes approximated by immunohistochemistry were analyzed in 633 patients with invasive breast carcinoma (excluding those with HER2 targeted therapy). Significant independent predictors for recurrence included: high histologic grade (p = 0.004), presence of lymphovascular invasion (p = 0.004), fibrotic focus (p = 0.020), mild lymphocytic infiltrate (p = 0.013), high TNM stage (p < 0.001), and HER2-overexpressing (p = 0.004) and basal-like (p < 0.001) molecular subtypes. A morphologic-molecular recurrence predictive model based on these features was useful in recurrence prediction, independent of treatment modalities, and was able to refine the traditional prognostic tools of histologic grade, TNM stage, and Nottingham prognostic index, particularly for intermediate-risk groups, and to refine the luminal group molecular subtypes. Such findings were reproducible with a validation cohort. CONCLUSION: TNM stage, histologic grade, lymphovascular invasion, fibrotic focus, mild lymphocytic infiltrate, HER2-overexpressing and basal-like molecular subtypes were important independent recurrence risk factors for breast cancer. This morphologic-molecular model was robust in recurrence prediction and refined recurrence risk stratified by the traditional prognostic parameters, independent of treatment modalities.

Androgen receptor expression shows distinctive significance in ER positive and negative breast cancers.

BACKGROUND: Androgen receptor (AR), a nuclear steroid hormone receptor, is differentially expressed in breast cancer subgroups with distinct clinical implications. METHODS: To investigate the clinical significance of AR in breast cancers more precisely, the expression of AR in a large cohort of breast cancer was correlated with clinicopathological features, biomarker expression, and patients' survival according to different molecular groupings in this study. RESULTS: Higher AR expression was found in ER+ (57.8 %) than in ER- (24.7 %) cancers. In the ER+ cancers, AR expression was associated with favorable clinicopathological features, including lower grade (p < .001), lower pT stage (p < .001), and positivity for PR (p < .001). It was an independent prognostic factor for longer disease-free survival, mainly in the HER2+ luminal B cancers (hazard ratio [HR] = 0.251, 95 % CI 0.065-0.972, p = .045). In ER- cancers, AR expression was associated with features distinct from basal-like breast cancer, and such features were found in molecular apocrine (MA) cancers. AR correlated with presence of extensive in situ component (p = .006) and apocrine phenotype (p < .001), HER2 (p = .026), and EGFR (p = .048), but negatively with c-kit (p = .041), CK5/6 (p < .001), CK14 (p = .002), and αB-crystallin (p = .038). However, AR expression was found only in 37.8 % of immunohistochemically defined MA. Of note, AR-MA appeared to have a trend of worse overall survival than AR+MA. CONCLUSIONS: AR expression was different in ER+ and ER- cancers and had different clinical implications. AR alone may not be a good marker for MA subtype. Its expression in MA may have substantial prognostic implication and as such warrants further validation.

TTF-1 expression in breast carcinoma: an unusual but real phenomenon.

AIMS: To evaluate thyroid transcription factor-1 (TTF-1) expression in a large cohort of invasive breast carcinomas (IBCs) using two commercially available monoclonal antibody clones (8G7G3/1 and SPT24). METHODS AND RESULTS: Nuclear and cytoplasmic TTF-1 expression was evaluated in 1132 primary IBCs and 208 primary pulmonary carcinomas using tissue microarray (TMA) sections. TTF-1 nuclear expression was detected in one of 1132 (0.09%) IBCs by 8G7G3/1. In pulmonary carcinoma, TTF-1 expression was detected in 149 (71.6%) and 147 (70.6%) cases by 8G7G3/1 and SPT24, respectively, with no significant difference being seen between the two clones (P = 0.839), and there was good consistency between these two antibodies in differentiating breast and pulmonary carcinomas (kappa value 0.905; P < 0.001). Both clones showed high specificity but relatively low sensitivity. Cytoplasmic TTF-1 expression was detected in 44 IBCs by 8G7G3/1, and this particular expression pattern was an independent adverse prognostic factor. CONCLUSIONS: Both TTF-1 antibodies (clones 8G7G3/1 and SPT24) were useful in differentiating breast from pulmonary carcinomas. Nuclear expression of TTF-1 was detected in IBCs by 8G7G3/1, but not by SPT24. Cytoplasmic expression of 8G7G3/1 was seen in IBC for the first time, and was an independent unfavourable prognostic factor.