Lymphangiosis carcinomatosa in squamous cell carcinomas of larynx and hypopharynx – value of conventional evaluation and additional immunohistochemical staining of D2-40

<p>Abstract</p> <p>Background</p> <p>Recent studies revealed a predictive value of lymphatic vessel invasion (L1) for the nodal metastasizing and poor prognosis in malignant tumors at different sites. The monoclonal antibody D2-40 (podoplanin) stains specifically endothelial cells of lymphatic vessels and improves the search for L1. However, the importance of this immunohistochemical staining was not investigated in squamous cell carcinomas (SCC) of larynx and hypopharynx.</p> <p>Aim</p> <p>This study was performed to compare the diagnostic potential of convential and immunohistochemical determination of L1 in SCC of larynx and hypopharynx with special respect to the predictive value for nodal metastasizing and prognosis.</p> <p>Methods</p> <p>119 SCCs of the larynx (n = 70) respectively hypopharynx (n = 49) were investigated. The lymphatic vessel invasion was assessed by conventional method (HE stain) and immunohistochemical staining with an antibody against D2-40 (DAKO, Germany). Immunohistochemistry was performed in accordance with manufacturer's protocol. L1 was searched microscopically in a standardized magnification (×200) in serial sections of tumor samples (1 section per cm tumor diameter).</p> <p>Results</p> <p>The immunohistochemical investigation did not show significant advantages for the prediction of regional nodal metastases. Despite a low sensitivity (< 50%) in both methods, the specifity can reach 80%. The negative predictive value in both methods seems acceptable (up to 80%), whereas the positive predictive value is not higher than 64%. Cases with L1 detected either conventionally or immunohistochemically did not show a significant shorter survival than cases with L0. However, a non-significant shorter survival was found. Only in SCC of hypopharynx, a combination of both methods revealed patients with a significant worse prognosis.</p> <p>Conclusion</p> <p>The status of lymphatic vessel invasion should be documented in standardized tumor reports. A benefit of an additional immunohistochemical investigation was not found, for the daily routine HE-stain seems sufficient.</p

Stop! In the name of transforming growth factor-β: keeping estrogen receptor-α-positive mammary epithelial cells from proliferating

Recent genetic and cell biological studies illustrate the importance of active transforming growth factor-β signaling in preventing the proliferation of estrogen receptor-positive cells in the normal mammary gland, and suggest how the loss of this inhibition may be important in early breast cancer progression

The expression of the ubiquitin ligase subunit Cks1 in human breast cancer

INTRODUCTION: Loss of the cell-cycle inhibitory protein p27(Kip1 )is associated with a poor prognosis in breast cancer. The decrease in the levels of this protein is the result of increased proteasome-dependent degradation, mediated and rate-limited by its specific ubiquitin ligase subunits S-phase kinase protein 2 (Skp2) and cyclin-dependent kinase subunit 1 (Cks1). Skp2 was recently found to be overexpressed in breast cancers, but the role of Cks1 in these cancers is unknown. The present study was undertaken to examine the role of Cks1 expression in breast cancer and its relation to p27(Kip1 )and Skp2 expression and to tumor aggressiveness. METHODS: The expressions of Cks1, Skp2, and p27(Kip1 )were examined immunohistochemically on formalin-fixed, paraffin-wax-embedded tissue sections from 50 patients with breast cancer and by immunoblot analysis on breast cancer cell lines. The relation between Cks1 levels and patients' clinical and histological parameters were examined by Cox regression and the Kaplan–Meier method. RESULTS: The expression of Cks1 was strongly associated with Skp2 expression (r = 0.477; P = 0.001) and inversely with p27(Kip1 )(r = -0.726; P < 0.0001). Overexpression of Cks1 was associated with loss of tumor differentiation, young age, lack of expression of estrogen receptors and of progesterone receptors, and decreased disease-free (P = 0.0007) and overall (P = 0.041) survival. In addition, Cks1 and Skp2 expression were increased by estradiol in estrogen-dependent cell lines but were down-regulated by tamoxifen. CONCLUSION: These results suggest that Cks1 is involved in p27(Kip1 )down-regulation and may have an important role in the development of aggressive tumor behavior in breast cancer

Essential function for ErbB3 in breast cancer proliferation

The overexpression of the ErbB family of tyrosine kinase receptors is thought to be important in the development of many breast tumours. To date, most attention has focused on the ErbB2 receptor. Now, in a recent report, it has been shown that ErbB3 is a critical partner for the transforming activity of ErbB2 in breast cancer cells. Importantly, the proliferative signals from this transforming complex appear to act via the PI-3 kinase pathway

Tamoxifen may prevent both ER+ and ER- breast cancers and select for ER- carcinogenesis: an alternative hypothesis

INTRODUCTION: Breast Cancer Prevention Trial (BCPT) and Multiple Outcomes of Raloxifene (MORE) data have been interpreted to indicate that tamoxifen reduces the risk of ER+ but not ER- breast carcinogenesis. We explored whether these data also support an alternative hypothesis, that tamoxifen influences the natural history of both ER+ and ER- cancers, that it may be equally effective in abrogating or delaying ER- and ER+ carcinogenesis, and place selection pressure, in some cases, for the outgrowth of ER- cancers. METHODS: BCPT and MORE data were used to investigate whether: first, tamoxifen could reduce equally the emergence of ER- and ER+ tumors; and second, tamoxifen could select a fraction of emerging ER+ cancers and promote their transformation to ER- cancers. Assuming that some proportion, Z, of ER+ tumors becomes ER- after tamoxifen exposure and that the risk reduction for both ER- and ER+ tumors is equal, we solved for both the transformation rate and the risk reduction rate. RESULTS: If tamoxifen equally reduces the incidence of ER+ and ER- tumors by 60%, the BCPT results are achieved with a transformation of approximately Z = 20% of ER+ to ER- tumors. Validation with MORE data using an equal risk reduction of 60% associated with tamoxifen produces an almost identical transformation rate Z of 23%. CONCLUSION: Data support an alternative hypothesis that tamoxifen may promote ER- carcinogenesis from a precursor lesion that would otherwise have developed as ER+ without tamoxifen selection

Lymph vascular invasion in invasive mammary carcinomas identified by the endothelial lymphatic marker D2-40 is associated with other indicators of poor prognosis

<p>Abstract</p> <p>Background</p> <p>Immunohistochemical studies of lymphatic vessels have been limited by a lack of specific markers. Recently, the novel D2-40 antibody, which selectively marks endothelium of lymphatic vessels, was released. The aim of our study is to compare lymphatic and blood vessel invasion detected by hematoxylin and eosin (H&E) versus that detected by immunohistochemistry, relating them with morphologic and molecular prognostic factors.</p> <p>Methods</p> <p>We selected 123 cases of invasive mammary carcinomas stratified into three subgroups according to axillary lymph node status: macrometastases, micrometastases, and lymph node negative. Lymphatic (LVI) and blood (BVI) vessel invasion were evaluated by H&E and immunohistochemistry using the D2-40 and CD31 antibodies, and related to histologic tumor type and grade, estrogen and progesterone receptors, E-cadherin, Ki67, p53, and Her2/<it>neu </it>expression.</p> <p>Results</p> <p>LVI was detected in H&E-stained sections in 17/123 cases (13.8%), and in D2-40 sections in 35/123 cases (28.5%) (Kappa = 0.433). BVI was detected in H&E-stained sections in 5/123 cases (4.1%), and in CD31 stained sections in 19/123 cases (15.4%) (Kappa = 0.198). LVI is positively related to higher histologic grade (p = 0.013), higher Ki67 expression (p = 0.00013), and to the presence of macrometastases (p = 0.002), and inversely related to estrogen (p = 0.0016) and progesterone (p = 0.00017) receptors expression.</p> <p>Conclusion</p> <p>D2-40 is a reliable marker of lymphatic vessels and is a useful tool for lymphatic emboli identification in immunostained sections of breast carcinomas with higher identification rates than H&E. Lymphatic vessel invasion was related to other features (high combined histologic grade, high Ki67 score, negative hormone receptors expression) associated with worse prognosis, probable reflecting a potential for lymphatic metastatic spread and aggressive behavior.</p

Immunostaining with D2–40 improves evaluation of lymphovascular invasion, but may not predict sentinel lymph node status in early breast cancer

<p>Abstract</p> <p>Background</p> <p>Sentinel lymph node (SLN) biopsy is a widely used diagnostic procedure in the management of early breast cancer. When SLN is free of metastasis, complete axillary dissection may be skipped for staging in clinically N0 patients, allowing a more conservative procedure. Histological tumor features that could reliably predict SLN status have not yet been established. Since the degree of tumor lymphangiogenesis and vascularization may theoretically be related to the risk of lymph node metastasis, we sought to evaluate the relationship between lymph vessel invasion (LVI), lymphatic microvascular density (LVD), microvascular density (MVD) and VEGF-A expression, with SLN status and other known adverse clinical risk factors.</p> <p>Methods</p> <p>Protein expression of D2–40, CD34, and VEGF-A was assessed by immunohistochemistry on paraffin-embedded sections of primary breast cancer specimens from 92 patients submitted to SLN investigation. The presence of LVI, the highest number of micro vessels stained for D2–40 and CD34, and the protein expression of VEGF-A were compared to SLN status, clinicopathological features and risk groups.</p> <p>Results</p> <p>LVI was detected in higher ratios by immunostaining with D2–40 (p < 0.0001), what would have changed the risk category from low to intermediate in four cases (4.3%). There was no association between LVI and other angiogenic parameters determined by immunohistochemistry with SLN macrometastases, clinical features or risk categories.</p> <p>Conclusion</p> <p>Assessment of LVI in breast carcinoma may be significantly increased by immunostaining with D2–40, but the clinical relevance of altering the risk category using this parameter may not be advocated according to our results, neither can the use of LVI and LVD as predictors of SLN macrometastasis in early breast cancer.</p

Co-Depletion of Cathepsin B and uPAR Induces G0/G1 Arrest in Glioma via FOXO3a Mediated p27Kip1 Upregulation

Cathepsin B and urokinase plasminogen activator receptor (uPAR) are both known to be overexpressed in gliomas. Our previous work and that of others strongly suggest a relationship between the infiltrative phenotype of glioma and the expression of cathepsin B and uPAR. Though their role in migration and adhesion are well studied the effect of these molecules on cell cycle progression has not been thoroughly examined.Cathepsin B and uPAR single and bicistronic siRNA plasmids were used to downregulate these molecules in SNB19 and U251 glioma cells. FACS analysis and BrdU incorporation assay demonstrated G0/G1 arrest and decreased proliferation with the treatments, respectively. Immunoblot and immunocyto analysis demonstrated increased expression of p27(Kip1) and its nuclear localization with the knockdown of cathepsin B and uPAR. These effects could be mediated by alphaVbeta3/PI3K/AKT/FOXO pathway as observed by the decreased alphaVbeta3 expression, PI3K and AKT phosphorylation accompanied by elevated FOXO3a levels. These results were further confirmed with the increased expression of p27(Kip1) and FOXO3a when treated with Ly294002 (10 microM) and increased luciferase expression with the siRNA and Ly294002 treatments when the FOXO binding promoter region of p27(Kip1) was used. Our treatment also reduced the expression of cyclin D1, cyclin D2, p-Rb and cyclin E while the expression of Cdk2 was unaffected. Of note, the Cdk2-cyclin E complex formation was reduced significantly.Our study indicates that cathepsin B and uPAR knockdown induces G0/G1 arrest by modulating the PI3K/AKT signaling pathway and further increases expression of p27(Kip1) accompanied by the binding of FOXO3a to its promoter. Taken together, our findings provide molecular mechanism for the G0/G1 arrest induced by the downregulation of cathepsin B and uPAR in SNB19 and U251 glioma cells