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Abstract: One of the hotspots in clinical research today is molecular genetic analysis of structural DNA alterations. Working with DNA obtained from formalin-fixed and paraffin-embedded (FFPE) tissue specimens is particularly challenging, due to cross-linking and fragmentation of DNA. We performed a comparative analysis of DNA extraction methods from FFPE tissue using two in-house protocols and Qiagen (QIAamp DNA FFPE Tissue Kit) and Roche (High Pure FFPET DNA Isolation Kit) commercial kits presented in the Russian market and used in clinical practice. To assess the quantity and quality of the isolated DNA, we used the real-time PCR to rate DNA yield, the inhibited impurity content and the degree of fragmentation. Our findings may be useful for a medical laboratory that performs testing of somatic mutations for the targeted therapy selection, and researchers who specialized in tumor genome structure studies in respect to patient prognosis and prediction of the sensitivity or resistance of tumor cells to therapy. Keywords: DNA extraction, formalin-fixed and paraffin-embedded (FFPE) tissue blocks, QIAamp DNA FFPE Tissue Kit, High Pure FFPET DNA Isolation Kit, PCR.Download Full Article |
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Abstract: Introduction: In this study we evaluate the relationship of PSA and PSAD with the degree of Gleason’s score of prostate cancer in transrectal ultrasound guided biopsy specimens. Methods: From March 2003 to October 2009, 1025 transractal ultrasound guided biopsies were performed in our hospital. PSA was measured by monoclonal antibody method and PSAD was calculated. The Gleason grade of the detected tumors in the biopsy specimens was classified as low, moderate and high grade. Data were analyzed by SPSS software. Results: 292 patients were diagnosed to have prostate adenocarcinoma. There was an acceptable correlation between PSA (P=0.001) and PSAD of the specimens (P = 0.013) with Gleason grades. PSA level showed a statistically significant difference between the low and high grade groups (P=0.005) and the intermediate and high grade groups (p=0.014). A statistically significant difference of PSAD level was seen only between the low and high grade (P=0.006) groups Conclusions: PSA and PSAD are both effective diagnostic tools for detection of prostate cancer; PSA level has a valuable role in predicting Gleason pattern higher than 7/10 and it can be the predictor of advanced pathological features but PSAD is effective in prediction of Gleason pattern lower than 5/10. Keywords: Prostate specific antigen, Prostate cancer, Gleason’s score, Tumor Grading, Gleason Grading.Download Full Article |
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Abstract: Epithelial Mesenchymal Transition (EMT) is the transdifferentiation of epithelial cells into a mesenchymal phenotype. This process occurs during embryogenesis but also in wound healing and in tumors. The neoplastic EMT is characterized by variably complete shedding of epithelial architectural features and acquisition of mesenchymal traits. In immunohistochemistry a variable coexpression of cytokeratins, vimentin or alpha-smooth muscle actin with loss of E-cadherin and other interepithelial adhesion molecules is characteristic. Such transition is associated with mutations both at the genetic (somatic) and epigenetic levels and is believed to confer a more advantageous phenotype for local and distant spread of cancer cells. Mammary carcinoma can exhibit EMT features in humans and mice and it tends to occur more frequently in women with tumors bearing a worse prognosis such as the claudin low subtype within the triple negative cancer. Missense mutation of TP53 is one of the most common mutations in cancer and it is frequently found in EMT tumor types, often with a more aggressive behavior. The current literature and survey of our mouse EMT cases in the Genomic Pathology Center image archives demonstrate a synergy between p53 and EMT that is independent of the initiating oncogene. However, p53 mutation is not sufficient or causal for EMT. Moreover, despite the local malignant behavior, processes such as spontaneous metastases and Mesenchymal Epithelial Transition (MET) appear not to be as frequent and obvious as previously hypothesized. Keywords: p53, EMT, metastasis, MET, breast cancer, mouse model, triple negative, claudin low.Download Full Article |
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Abstract: Objectives:Carcinoembryonic antigen (CEA), epidermal growth factor (EGF) and leptin have been reported to be intimately intertwined in lung carcinogenesis.However, few studies have simultaneously examined these proteins in lung cancer and whether a correlation exist among them remains unclear. Here, we compared the levels of CEA, EGF and leptin in non-small-cell lung cancer (NSCLC) patients and controls and evaluated the possible associations among them. Methods:97 patients ranged from 30 to 83 years of age were studied. Serum CEA, EGF and leptin levels were determined following a standard protocol. The relationships between these proteins and clinicopathological factors were evaluated by Wilcoxon rank sum or Kruskal-Wallis H test. Spearman rank-correlation were used to determine the correlations among CEA, EGF and leptin. Co-expression of these proteins in NSCLC tissues was examined by immunofluorescence. Results: Serum CEA and leptin levels in NSCLC patients were significantly higher compared to controls (both P = 0.000), but no statistically significant difference was found for EGF. CEA and EGF were not associated with the tumor-related factors, but leptin was strongly correlated with sex (P = 0.005). Significant correlations among these proteins were found when the patients were categorized into subgroups. Co-expresstion of these proteins was significantly enhanced with lung carcinogenesis. Conclusions:CEA, EGF and leptin may interplay and play vital roles in the pathogenesis of NSCLC. Besides CEA, the leptin levels were also significantly higher in NSCLC patients than in controls. Determination of preoperative leptin levels may prove useful for screening and predicting NSCLC. Keywords: Non-small-cell lung cancer, Carcinoembryonic antigen (CEA), Epidermal growth factor (EGF), Leptin,Immunofluorescence,Correlation.Download Full Article |
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Abstract: Histopathology, the current “gold standard”, is prone to human errors as it depends on expert interpretation of the microscopically derived cellular and sub-cellular information for tissue diagnosis. Further, this light microscope based approach requires preparation of appropriately stained specimens of micro-thin tissue sections from the formalin-fixed and paraffin-embedded (FFPE) blocks of tissue samples. We report a method that provides quantitative feedback about tissue diagnosis by measuring depth-sensitive Raman spectra from the intact FFPE tissue blocks without requiring preparation of any thin tissue sections or any other processing. The FFPE blocks of pathologically certified cancerous and normal breast tissues were used for validating the approach. The measured depth-sensitive Raman spectra were mathematically de-paraffinized for retrieving the characteristic tissue Raman signatures using scaled-subtraction. A multivariate analysis of the scaled-subtracted, depth-sensitive Raman spectra employing a probability-based diagnostic algorithm developed using the framework of sparse multinomial logistic regression (SMLR) provided a sensitivity and specificity of up to 100% towards cancer based on leave-one-block-out cross validation. The results of this exploratory study suggest that depth-sensitive Raman spectroscopy along with a multivariate statistical algorithm can provide a valuable alternate diagnostic modality in clinical pathology setting for discriminating cancerous from normal FFPE tissue blocks. Keywords: Formalin-fixed and paraffin-embedded (FFPE) tissue blocks, Depth-sensitive Raman spectroscopy, Scale-subtraction.Download Full Article |