Direct Comparison of Metastasis-Related miRNAs Expression Levels in Circulating Tumor Cells, Corresponding Plasma, and Primary Tumors of Breast Cancer Patients.

BACKGROUND: Circulating tumor cells (CTCs) and microRNAs (miRNAs) are important in liquid biopsies in which peripheral blood is used to characterize the evolution of solid tumors. We evaluated the expression levels of miR-21, miR-146a, miR-200c, and miR-210 in CTCs of breast cancer patients with verified metastasis and compared their expression levels in corresponding plasma and primary tumors. METHODS: Expression levels of the miRNAs were quantified by quantitative reverse transcription PCR (RT-qPCR) in (a) 89 primary breast tumors and 30 noncancerous breast tissues and (b) CTCs and corresponding plasma of 55 patients with metastatic breast cancer and 20 healthy donors. For 30 of these patients, CTCs, corresponding plasma, and primary tumor tissues were available. RESULTS: In formalin-fixed, paraffin-embedded tissues, these miRNAs were differentially expressed between primary breast tumors and noncancerous breast tissues. miR-21 (P < 0.001) and miR-146a (P = 0.001) were overexpressed, whereas miR-200c (P = 0.004) and miR-210 (P = 0.002) were underexpressed. In multivariate analysis, miR-146a overexpression was significantly [hazard ratio 2.969 (1.231-7.157), P = 0.015] associated with progression-free survival. In peripheral blood, all miRNAs studied were overexpressed in both CTC and corresponding plasma. There was a significant association between miR-21 expression levels in CTCs and plasma for 36 of 55 samples (P = 0.008). In plasma, ROC curve analysis revealed that miR-21, miR-146a, and miR-210 could discriminate patients from healthy individuals. CONCLUSIONS: Metastasis-related miRNAs are overexpressed in CTCs and corresponding plasma; miR-21 expression levels highly correlate in CTCs and plasma; and miR-21, miR-146a, and miR-210 are valuable plasma biomarkers for discriminating patients from healthy individuals.

Expression of EMT inducers integrin-linked kinase (ILK) and ZEB1 in phyllodes breast tumors is associated with aggressive phenotype.

Phyllodes tumors (PTs) of the breast constitute an uncommon group of mammary fibroepithelial lesions with ambiguous biologic behavior. Recent evidence suggests that epithelial mesenchymal transition (EMT), a driving force of cancer progression is implicated in PTs pathogenesis. Integrin-linked kinase (ILK), a focal adhesion kinase, has been implicated in cancer and EMT and represents a novel cancer therapeutic target. In this study, we aimed to investigate ILK and EMT markers expression in phyllodes breast tumors in relation to tumor grade. Expression of ILK and EMT markers E-cadherin, ß-catenin, Ν-cadherin, vimentin, Snail, ZEB1 and Twist was evaluated by immunohistochemistry in paraffin-embedded tissue sections from 96 human phyllodes breast tumors (48 benign, 27 borderline, 21 malignant). Cytoplasmic and nuclear immunopositivity of ILK were observed in both the epithelial and the stromal component of phyllodes breast tumors and were significantly higher with increasing tumor grade. An EMT-related expression profile consisting of decreased membranous and increased nuclear/cytoplasmic immunoreactivity of E-cadherin and ß-catenin and increased expression of N-cadherin, vimentin, Snail, ZEB1 and Twist was observed in tumor epithelial and stromal component and was significantly associated with malignant phyllodes breast tumor histopathology. Interestingly, there was a significant correlation of ILK expression with all of the EMT markers examined. Our results suggest that EMT significantly contributes to phyllodes tumor pathogenesis and originally implicate ILK and ZEB1 in phyllodes tumors malignant phenotype.

Tumour expression of lymphangiogenic growth factors but not lymphatic vessel density is implicated in human cervical cancer progression.

AIMS: The aim of the study was to investigate the role of lymphangiogenesis in human cervical cancer progression. METHODS: The expression of VEGF-C, VEGF-D, VEGFR-3, podoplanin (D2-40), LYVE-1 and Prox-1 was studied by immunohistochemistry in 72 cases of invasive squamous cell carcinoma of the uterine cervix. For lymphatic endothelial markers lymphatic vessel density (LVD) was assessed. Correlations with lymphatic vessel invasion, nodal metastases, tumour grade, FIGO stage, and inflammation were also evaluated. RESULTS: VEGF-D expression significantly correlated with lymph node metastasis, lymphatic emboli and FIGO stage as well as with peritumoural LVD. A marginally significant correlation was also found between the expression of VEGF-C and prognostic parameters. Lymphatic tumour emboli were successfully identified using D2-40 immunohistochemistry and peritumoural D2-40 LVD significantly correlated with lymphatic vessel invasion. However, LVD as assessed by multiple lymphatic markers was not associated with lymphatic metastasis. There was a significant correlation of Prox-1 and LYVE-1 LVD with the inflammatory stromal reaction. CONCLUSIONS: Although LVD as assessed by multiple lymphatic markers was not correlated with prognostic parameters, tumour expression of lymphangiogenic growth factors seems to be critically implicated in lymphatic metastasis and cervical carcinoma progression.