Flow Cytometric Assessment of Endothelial and Platelet Microparticles in Patients With Atrial Fibrillation Treated With Dabigatran.

The prothrombotic state in patients with atrial fibrillation (AF) is related to endothelial injury, the activation of platelets and the coagulation cascade. We evaluated the levels of platelet- (CD42b) and endothelial-derived (CD144) microparticles in the plasma patients with non-valvular AF treated with dabigatran at the time of expected minimum and maximum drug plasma concentrations. Following that, we determined the peak dabigatran plasma concentration (cpeak ). CD42b increased after taking dabigatran (median [IQR] 36.7 [29.4-53.3] vs. 45.6 [32.3-59.5] cells/µL; p = 0.025). The concentration of dabigatran correlated negatively with the post-dabigatran change in CD42b (ΔCD42b, r = -0.47, p = 0.021). In the multivariate model, the independent predictors of ΔCD42b were: cpeak (HR -0.55; with a 95% confidence interval, CI [-0.93, -0.16]; p = 0.007), coronary artery disease (CAD) (HR -0.41; 95% CI [-0.79, -0.02]; p = 0.037) and peripheral artery disease (PAD) (HR 0.42; 95% CI [0.07, 0.74]; p = 0.019). CD144 did not increase after dabigatran administration. These data suggest that low concentrations of dabigatran may be associated with platelet activation. PAD and CAD have distinct effects on CD42b levels during dabigatran treatment.

Brief fixation enables same-day breast cancer diagnosis with reliable assessment of hormone receptors, E-cadherin and HER2/Neu.

AIMS: Preoperative core needle biopsy (CNB) is commonly used to confirm the diagnosis of breast cancer. For treatment purposes and for determining histological type, especially in case of neoadjuvant therapy, oestrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) status and E-cadherin assessments are crucial. Considering the increasing demand for same-day diagnosis of breast lesions, an accelerated method of CNB processing was developed, in which the tissue fixation time is radically reduced. METHODS: To determine whether short fixation time frustrates assessment of ER, PR and E-cadherin immunohistochemistry (IHC) and HER2 fluorescence in situ hybridisation (FISH), 69 consecutive patients with 70 invasive breast carcinomas were included through the same-day diagnostics programme of breast lesions of the Radboud university medical center and the hospital Pantein. IHC for ER, PR and E-cadherin and HER2 FISH were compared between CNBs fixed for approximately 60-90 min and traditionally fixed resection specimens. RESULTS: Overall agreement between CNBs and resection specimens was 98.6% for ER (p<0.001; κ=0.93), 90.0% for PR (p<0.001; κ=0.75), 100% for E-cadherin (p<0001; κ=1.00) and 98.6% (p<0.001; κ=0.94) for HER2 FISH. Positive and negative predictive values were respectively 98.4% and 100% for ER, 95.9% and 76.2% for PR, 100% and 100% for E-cadherin and 90% and 100% for HER2 FISH. CONCLUSIONS: Hormone receptors and E-cadherin IHC and HER2 FISH are highly comparable between briefly fixed CNBs and the corresponding traditionally fixed resection specimens, and can therefore reliably be used in the daily clinical practice of same-day diagnostics of breast cancer.

Correlation between invasion mode and the histologic risk assessment model in oral squamous cell carcinoma.

PURPOSE: The histologic risk assessment (HRA) model was proposed to assess clinical outcome of oral squamous cell carcinomas (SCCs), and its prognostic value has been confirmed in several studies, but its underlying molecular mechanisms has not been fully investigated. The objective of this study was to evaluate the association between immunohistochemical invasion mode of head and neck SCC and the HRA grading system. METHODS: The HRA model consisting of risk scores (RSs), based on perineural invasion (PI), lymphocytic infiltration (LI), and worst pattern of invasion (WPOI), was used for grading 80 samples of head and neck SCC, followed by immunohistochemical staining with antibodies against E-cadherin, N-cadherin, and podoplanin. The two major invasion modes were considered as epithelial-mesenchymal-transition (EMT) and collective cell invasion (CCI) with profiles of E-cadherin-/N-cadherin+/podoplanin- and E-cadherin+/N-cadherin-/podoplanin+, respectively. All other immunohistochemical profiles were classified as partial/incomplete EMT. Statistical analysis was performed by Kruskal-Wallis and Mann-Whitney U tests (P < 0.05). RESULTS: Invasion pattern was significantly different among the three RSs (P = 0.01) and across the different scores of LI (P = 0.03) but not perineural invasion (PNI) (P = 0.07) and WPOI (P = 0.70). CONCLUSIONS: Based on our results, it seems that there is a difference in EMT, CCI, and partial/incomplete EMT among the variables of the HRA model which might help clarify its functioning system.

Detailed assessment of microvasculature markers in non-small cell lung cancer reveals potentially clinically relevant characteristics.

Tumor angiogenesis is important for the progression of cancer and is orchestrated by various factors associated with tumor vessels, tumor cells, and stromal cells. Angiogenic signaling in non-small cell lung cancer (NSCLC) needs to be further clarified, especially regarding existing and upcoming therapeutic approaches. Expression of CD34, CD105, Mel-CAM, VE-cadherin, D2-40, VEGF, VEGFR1, and VEGFR2 was assessed immunohistochemically on a cohort of 371 well documented, surgically resected NSCLC using a standardized tissue microarray platform. Extensive clinical data and a postoperative follow-up period of up to 18 years allowed us to assess clinicopathological correlations in detail. Microvasculature in NSCLC was significantly denser at the tumor periphery as compared to the tumor center. Squamous cell carcinomas (SCC) were associated with a notably lower microvessel density (MVD) than adenocarcinomas (ACA). CD105 was present at significantly higher levels on stromal cells of ACA as compared to SCC. Expression of VE-cadherin by tumor cells (6% of cases, mainly ACA) as well as decreased MVD in the tumor centers was independently associated with poor prognosis in the entire cohort. Low MVD in SCC might be related to lower efficacy of and fatal bleeding during therapy with bevacizumab. In other NSCLC entities for which treatment with VEGF inhibitors is studied in clinical trials, the predictive value of MVD for therapy response merits to be prospectively examined. Our data suggest that patients with ACA may be candidates for therapies targeting CD105. VE-cadherin is another promising target for therapy, but its expression also provides independent prognostic information.

Immunohistochemical assessment of CD1a-positive Langerhans cells and their relationship with E-cadherin in minor salivary gland tumors.

OBJECTIVE: The aim of this study was to investigate the presence of CD1a-positive Langerhans cells and their relationship with E-cadherin in minor salivary gland tumors. METHODS: Twenty-seven minor salivary gland tumors were investigated using immunohistochemistry for CD1a and E-cadherin. RESULTS: A significant difference regarding the mean density of CD1a-positive Langerhans cells was observed between pleomorphic adenomas and malignant tumors studied (P = 0.001). No CD1a-positive cells were detected in most cases (n = 5) of cystic adenoid carcinomas. CD1a-positive cells were detected in one mucoepidermoid carcinoma case, and six low-grade polymorphous adenocarcinomas cases. Comparison of the mean density of CD1a-positive cells between the three malignant tumors showed no significant difference (P = 0.127). No significant difference was observed in the presence of E-cadherin between tumors (P = 0.73), but it was detected in 24 cases. CONCLUSIONS: The lack of CD1a-positive in malignant salivary gland tumors facilitates the neoplastic development and suggests that these cells might be useful as auxiliary diagnostic and prognostic tool in minor salivary gland tumors. Furthermore, it is suggested that E-cadherin mediates cell adhesion in these tumors although we did not demonstrate significance.

Combined assessment of EGFR pathway-related molecular markers and prognosis of NSCLC patients.

The purpose of this study is to evaluate the prognostic value of the combined assessment of multiple molecular markers related to the epidermal growth factor receptor (EGFR) pathway in resected non-small cell lung cancer (NSCLC) patients. Tumour specimens of 178 NSCLC patients were collected and analysed for EGFR and KRAS mutation status by DNA sequencing, and for EGFR copy number by fluorescent in situ hybridisation. Tissue microarrays were generated and used to determine the expression of multiple EGFR pathway-related proteins by immunohistochemistry. We analysed the association between each marker and patient prognosis. Univariate analyses for each clinical variable and each molecular marker were performed using Kaplan-Meier curves and log-rank tests. From these results, we selected the variables KRAS mutations and expression of cytoplasmic EGFR, granular pERK, nuclear pSTAT3, cytoplasmic E-cadherin and cytoplasmic pCMET to enter into a Cox proportional hazards model, along with stage as the strongest clinical variable related with prognosis. Of the EGFR-related markers evaluated here, the markers EGFR, pERK, pSTAT3, E-cadherin, pCMET and mutations in KRAS were associated with survival when analysed in combination in our patient cohort, with P=0.00015 as the P-value for a test of the additional impact of markers on prognosis, after taking stage into consideration. Confirmation of the impact of these markers in independent studies will be necessary.

Immunohistochemical assessment of E-cadherin and beta-catenin in trichofolliculomas and trichoepitheliomas.

BACKGROUND: Trichofolliculomas and trichoepitheliomas are benign skin neoplasms originating from hair follicle cells. They result from defects in the signaling pathways that regulate hair follicle morphogenesis and regeneration. Thus they seem to be an excellent model of these processes. It is known that the E-cadherin/beta-catenin system of adhesion molecules plays a crucial role in the maintenance of tissue architecture. AIM: The aim of the present study was to investigate their involvement in benign hair follicle tumor development. METHODS: Semiquantitative intensity of expression were examined in formalin-fixed and paraffin-embedded tissue sections of 53 trichoepitheliomas, 15 trichofolliculomas and 19 normal skin samples by indirect immunohistochemistry. RESULTS: The intensity of E-cadherin/beta-catenin expression in tumor cells did not differ from controls. However, normal hair follicles cells exhibited membranous E-cadherin/beta-catenin expression, whereas both types of tumors, particularly trichoepitheliomas, showed E-cadherin/beta-catenin expression with a predominantly cytoplasmic localization. CONCLUSIONS: We suggest that this dystopic distribution of the E-cadherin/beta-catenin complex in hair follicle tumor cells may be a marker of cell-cell adhesion disruption which may contribute to the tumor formation.

Assessment of staging, prognosis and mortality of colorectal cancer by tumor markers: receptor erbB-2 and cadherins

OBJETIVO: Avaliação da expressão tumoral das proteínas c-erbB-2 e E-caderina e sua relação com o prognóstico, estadiamento e grau de diferenciação celular, em doentes com câncer colo-retal . MÉTODOS: O estudo incluiu 117 doentes com média de idade de 63.1 anos e com acompanhamento médio de 28.1 meses. O intervalo livre de doença, sobrevida, índice de recidiva e mortalidade específica foram os parâmetros avaliados. Anticorpos anti-oncoproteína c-erbB-2 (Dako) foram utilizados pela técnica da estreptavidiva-biotina. Considerou-se como positiva a presença desta proteína quando mais de 10% das células tumorais estivessem coradas. A proteína E-caderina foi estudada pelo anticorpo anti-E-caderina (Dako), sendo computada como positiva a amostra que apresentasse 50% ou mais das células coradas. A análise estatística utilizou o teste do qui-quadrado de Pearson, o teste exato de Fischer, a curva de Kaplan-Meier, o teste de log-rank e o teste de Wilcoxon ( variante de Breslow),sendo estabelecido nível de significância de 5%( p<0,05). RESULTADOS: 52 de 108 doentes estudados para c-erbB-2 foram positivos (48,1%), 47 de 93 doentes estudados para E-caderina foram negativos (50,5%). Estes dados não mostraram relação com estadiamento TNM (tumor, nódulo e metástase), com o grau de diferenciação celular e índice de recidiva tumoral. O intervalo livre de doença para os doentes positivos para c-erbB-2 e negativos para E-caderina foi de 68.0 meses e não diferiu daqueles que foram negativos para c-erbB-2 e positivos para E-caderina ( 55.0 meses - p = 0.5510). A sobrevida média para os doentes positivos para c-erbB-2 e negativos para E-caderina foi 75 meses sem diferença estatisticamente significante com o outro grupo de comparação( 61 meses - p = 0.5256). A mortalidade específica foi de 20.0% dos casos e não se correlacionou com a expressão do c-erbB-2 (p=0,446) ou da E-caderina(p=0,883). CONCLUSÃO: A expressão das proteínas c-erbB-2 e E-caderina em doentes portadores de adenocarcinoma colo-retal não apresentou correlação com o estadiamento e grau de diferenciação celular. Não houve da mesma forma relação com o prognóstico, no que diz respeito ao índice de recidiva da doença, intervalo livre de doença, sobrevida e mortalidade específica.

Assessment of staging, prognosis and mortality of colorectal cancer by tumor markers: receptor erbB-2 and cadherins.

PURPOSE: To evaluate the prognostic significance and correlation with staging and degree of cell differentiation of the tumoral expression of the proteins c-erbB-2 and E-cadherin, in patients with colorectal adenocarcinoma. METHODS: The study included 117 patients with an average age of 63.1 years and an average follow-up duration of 28.1 months. The disease-free interval, survival, incidence of recurrence and specific mortality were evaluated. c-erbB-2 anti-oncoprotein antibodies (Dako) were utilized via the streptavidin-biotin technique. Samples were considered to be positive for c-erbB-2 if 10% or more of the tumor cell membranes were stained. The anti-E-cadherin antibodies (Dako), evaluated this protein and is considered positive, if 50% or more of the cell membranes were stained. Statistical analysis was performed using Pearson's chi-squared test, Fisher's exact test, Kaplan-Meier's estimator, the log-rank test and Wilcoxon's test (Breslow version), setting the level of statistical significance at 5% (p<0.05). RESULTS: 52 of 108 patients studied for c-erbB-2 were positive (48.1%), 47 of 93 patients studied for E-cadherin were negative (50.5%). These data do not express any correlation with TNM (tumor, node and metastasis) staging and the degree of cell differentiation or with the tumor recurrence rate. The disease-free interval among patients who were positive for c-erbB-2 and negative for E-cadherin was 68.0 months and did not differ from those with c-erbB-2 negative and E-cadherin positive (55.0 months--p = 0.5510). The average survival among patients positive for c-erbB-2 and negative for E-cadherin was 75 months without statistical significance difference with the other group (61 months--p = 0.5256). Specific mortality occurred in 20.0% of the cases and did not correlate with the expression of c-erbB-2 (p=0.446), E-cadherin (p=0.883). CONCLUSION: The tumoral expression of c-erbB-2 and E-cadherin did not demonstrate a correlation with the staging and degree of cell differentiation, and it did not present prognostic value regarding disease recurrence, disease-free interval, survival and specific mortality among patients with colorectal adenocarcinoma.

Micrometastasis in the bone marrow of patients with lung cancer associated with a reduced expression of E-cadherin and beta-catenin: risk assessment by immunohistochemistry.

BACKGROUND: The presence of disseminated tumor cells in bone marrow is considered to be a premetastatic state, which is called micrometastasis. To evaluate the relationship between micrometastasis and cellular adhesion molecules in the primary lesion, E-cadherin and beta-catenin were immunohistochemically investigated. Methods. Fifty-eight patients with non-small cell lung cancer who underwent a complete resection were entered into this study. Tumor cells in bone marrow aspirates were detected by immunohistochemistry using cytokeratin (CK) 18. Immunohistochemical studies of E-cadherin and beta-catenin were performed in the corresponding primary tumor. RESULTS: CK-positive cells were detected in the bone marrow aspirates from 27 of 58 patients. A reduced expression of the E-cadherin and beta-catenin was found in 16 (27.6%) and in 22 (37.9%) of 58 patients, respectively. In 26 cases with a reduced expression of E-cadherin and/or beta-catenin, 16 cases had CK-positive cells, whereas 11 of 32 cases with normal expression of both factors had CK-positive cells (P=.0392). The patients with micrometastasis demonstrated an earlier recurrence (P =.0642) and a significantly poorer survival (P =.0437) than those without such cells. CONCLUSIONS: Micrometastasis in the bone marrow might be a significant predictor of poor prognosis, and a reduced expression of E-cadherin and beta-catenin are important determinants for the metastatic capability of individual cancer cells.