Integrin-Linked Kinase, Snail and Multidrug Resistance Protein 1: Three concordant players in the progression of non-small cell lung cancer.

BACKGROUND: Integrin Linked Kinase (ILK), Snail and Multidrug Resistance Protein 1 (MRP1) have been implicated in several cancers; however, their roles in non-small cell lung cancer (NSCLC) remain to be elucidated. AIM: Investigation of their expression in NSCLC tissue. Relationships among these proteins and their association with clinicopathological parameters were studied. MATERIALS AND METHODS: ILK, Snail and MRP1 expression were immunohistochemically assessed in 97 tumor tissues. Furthermore, western blot analysis for ILK, Snail and MRP1 in 6 cases of NSCLC was also performed. RESULTS: ILK overexpression, positive Snail and MRP1 expression were found in 46.4%, 36.1% and 49.5% of tumors respectively. ILK expression was significantly correlated with tumor grade (p=0.013), lymph node (LN) metastases (p=0.001) and stage (p=0.001). Positive Snail and MRP1 expression were significantly associated with LN metastasis (p=0.004 and 0.022, respectively) and advanced stage disease (p=0.018 and 0.024, respectively). MRP1 expression was significantly higher among adenocarcinoma cases compared to other types (p=0.001). ILK over-expression was significantly associated with up-regulation of Snail and MRP1 (p<0.001 both). Significant association was also, found between Snail and MRP1 expression (p=0.005). Moreover, the co-expression of two markers or more was significantly associated with less differentiation (p=0.011), advanced tumor status (p=0.030), LN metastasis (p<0.001) and advanced stage (p<0.001) disease. Western blot analysis validated immunohistochemical findings. CONCLUSION: ILK may have an important role in the progression of NSCLC, possibly through up-regulation of Snail and MRP1. ILK, Snail and MRP1 are important molecular markers for predicting carcinogenesis and progression of NSCLC.

Defective Beclin-1 and elevated hypoxia-inducible factor (HIF)-1α expression are closely linked to tumorigenesis, differentiation, and progression of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is an aggressive cancer with a poor prognosis. Autophagy and hypoxia have been involved in HCC tumorigenesis. In the present study, we examined the relationship between Beclin-1 expression and hypoxia-inducible factor (HIF)-1α expression in HCC by immunohistochemistry on 65 tumor specimens. Their correlations with clinicopathological features were also explored. There was a loss of Beclin-1 protein expression in 49.2 % of HCC. Beclin-1 expression was only significantly correlated with virus infection status (p = 0.025) and marginally associated with HCC grade (p = 0.057). Forty-two tumors (64.6 %) showed high HIF-1α expression, and it was significantly associated with large tumor size (p = 0.003), multifocal tumors (p = 0.038), and advanced stage (p = 0.043). Beclin-1 expression was significantly associated with HIF-1α expression (p = 0.001). HCC cases were further stratified according to their hypoxia status into hypoxic and normoxic groups. In the hypoxic group, Beclin-1 expression was negatively correlated with HCC high tumor grade (p < 0.001), advanced stage (p = 0.013), large size (p = 0.002), and multifocal tumors (p = 0.047). In the normoxic group, no significant relations between Beclin-1 expression and any of the clinicopathological parameters were identified. Our findings that reduced Beclin-1 and high HIF-1α expression are associated with the development and progression of HCC may provide molecular therapeutic targets toward inhibiting HCC development and progression.

Expression of Proteinase-activated Receptor-2 in the Esophageal Mucosa of Gastroesophageal Reflux Disease Patients: A Histomorphologic and Immunohistochemical Study.

Data are limited regarding the role of proteinase-activated receptor-2 (PAR-2) in the esophageal mucosa in gastroesophageal reflux disease (GERD) patients. Our aim was to study PAR-2 expression and its relationship with different GERD-related clinical and pathologic parameters. Histomorphologic alterations in eosophageal mucosa in nonerosive reflux disease (NERD) and erosive reflux disease (ERD) were also, evaluated. Endoscopic biopsies of the esophageal mucosa were obtained from 94 GERD patients and 20 participants for histopathologic analysis and PAR-2 immunohistochemical staining. The present study demonstrated significantly higher PAR-2 expression in GERD patients compared with control, whereas no significant differences were seen between NERD and ERD groups. PAR-2 expression significantly correlated with histologic score (r=0.572, P<0.001) and severity of heartburn (r=0.541, P<0.001). PAR-2 expression was significantly associated with basal cell hyperplasia, and dilated intercellular spaces and inflammatory cell count (P<0.05). Histologic analysis revealed GERD-related histomorphologic alterations in the esophageal mucosa of GERD patients with significant differences (P<0.05) among groups. Total histologic score was significantly correlated with heartburn (r=0.299, P=0.025) and endoscopic severity (r=0.359, P=0.027) in NERD and ERD patients, respectively. Taken together, this study provides evidence for the major role of PAR-2 in the pathogenesis of GERD and GERD-associated mucosal alterations.

Expression of a disintegrin and metalloprotease 8 and endostatin in human osteosarcoma: implication in tumor progression and prognosis.

BACKGROUND: A disintegrin and metalloprotease 8 (ADAM8) is a trans-membrane protein, which is involved in cell adhesion, signaling and migration as well as the proteolytic cleavage of various substrates. Endostatin is a potent inhibitor of angiogenesis. ADAM8 and Endostatin have been associated with multiple malignancies. However, their role in osteosarcoma is not fully elucidated. AIM: To determine the expression of ADAM8 and endostatin in osteosarcoma and to study their correlation with different clinicopathological parameters and patients' outcomes. MATERIAL AND METHODS: ADAM8 and endostatin expression were immunohistochemically evaluated in 61 primary osteosarcomas and 11 pulmonary metastatic osteosarcoma lesions. RESULTS: Among 61 primary osteosarcomas, ADAM8 was detected in 52 tumors (85.2%) and highly expressed in 33 cases (54.1%). Positive endostatin expression was found in 28 tumors (45.9%). Higher ADAM8 and decreased endostatin expression rates in metastatic lesions compared to primary osteosarcoma were found but these differences were not statistically significant (p=0.086 & 0.558 respectively). High ADAM8 expression score and positive endostatin expression were significantly correlated with tumor size, stage and distant metastasis (p<0.05). Survival analysis showed that high ADAM8 expression was associated with poor overall survival (OS) (p=0.0002). Multivariate analysis revealed that ADAM8 expression level was an independent prognostic parameter for the OS (p=0.017). CONCLUSION: Our data suggest that ADAM8 and endostatin play a role in osteosarcoma progression. High ADAM8 expression serves as a reliable marker for poor prognosis in osteosarcoma patients.

Clinicopathologic implications of EpCAM and Sox2 expression in breast cancer.

BACKGROUND: The purpose of this study was to investigate the clinicopathologic significance of EpCAM and Sox2 expression in breast cancer and to study their correlation during breast cancer progression. PATIENTS AND METHODS: EpCAm and Sox2 expression were assessed using immunohistochemistry in ductal carcinoma insitu (DCIS), invasive breast cancer (IBC) and matched lymph node metastasis (LNM), if present. RESULTS: EpCAM overexpression was found in 63.2% of DCIS, 72.2% of IBC and 74.4% of LNM. In IBC cases, EpCAM overexpression was associated with high grade (P < .001), large tumor size (P = .051), poor Nottingham Prognostic Index (NPI) (P = .006), histological tumor types (P = .044) and the triple negative phenotype (P = .008). LNM frequently reflected the expression phenotype of the matched primary tumors with no significant differences between LNM and their primary tumors (P = .564). Sox2 expression was detected in 47.4%, 33.3% and 54.7% of DCIS, IBC and LNM respectively. In DCIS group, Sox2 expression was significantly associated with comedo type (P = .037), negative ER (P = .012) and PR (P = .037) and the triple negative phenotype (P = .006). In IBC cases, Sox2 expression showed significant associations with high grade (P = .045), nodal spread (P = .037), poor NPI (P = .018) and the triple negative phenotype (P < .001). LNM showed significantly higher Sox2 expression rates than primary tumors (P < .001). Significant positive associations between EpCAM overexpression and Sox2 positivity in DCIS (P = .027), IBC (P = .001) and LNM (P < .001) were found. CONCLUSION: This study emphasized the potential role of EpCAM and Sox2 in breast carcinogenesis and revealed their involvement during breast cancer progression and LN metastases.

High-throughput protein expression analysis using tissue microarray technology of a large well-characterised series identifies biologically distinct classes of breast cancer confirming recent cDNA expression analyses.

Recent studies on gene molecular profiling using cDNA microarray in a relatively small series of breast cancer have identified biologically distinct groups with apparent clinical and prognostic relevance. The validation of such new taxonomies should be confirmed on larger series of cases prior to acceptance in clinical practice. The development of tissue microarray (TMA) technology provides methodology for high-throughput concomitant analyses of multiple proteins on large numbers of archival tumour samples. In our study, we have used immunohistochemistry techniques applied to TMA preparations of 1,076 cases of invasive breast cancer to study the combined protein expression profiles of a large panel of well-characterized commercially available biomarkers related to epithelial cell lineage, differentiation, hormone and growth factor receptors and gene products known to be altered in some forms of breast cancer. Using hierarchical clustering methodology, 5 groups with distinct patterns of protein expression were identified. A sixth group of only 4 cases was also identified but deemed too small for further detailed assessment. Further analysis of these clusters was performed using multiple layer perceptron (MLP)-artificial neural network (ANN) with a back propagation algorithm to identify key biomarkers driving the membership of each group. We have identified 2 large groups by their expression of luminal epithelial cell phenotypic characteristics, hormone receptors positivity, absence of basal epithelial phenotype characteristics and lack of c-erbB-2 protein overexpression. Two additional groups were characterized by high c-erbB-2 positivity and negative or weak hormone receptors expression but showed differences in MUC1 and E-cadherin expression. The final group was characterized by strong basal epithelial characteristics, p53 positivity, absent hormone receptors and weak to low luminal epithelial cytokeratin expression. In addition, we have identified significant differences between clusters identified in this series with respect to established prognostic factors including tumour grade, size and histologic tumour type as well as differences in patient outcomes. The different protein expression profiles identified in our study confirm the biologic heterogeneity of breast cancer and demonstrate the clinical relevance of classification in this manner. These observations could form the basis of revision of existing traditional classification systems for breast cancer.

Expression of luminal and basal cytokeratins in human breast carcinoma.

We have examined basal and luminal cell cytokeratin expression in 1944 cases of invasive breast carcinoma, using tissue microarray (TMA) technology, to determine the frequency of expression of each cytokeratin subtype, their relationships and prognostic relevance, if any. Expression was determined by immunocytochemistry staining using antibodies to the luminal cytokeratins (CKs) 7/8, 18 and 19 and the basal markers CK 5/6 and CK 14. Additionally, assessment of alpha-smooth muscle actin (SMA) and oestrogen receptor status (ER) was performed. The vast majority of the cases showed positivity for CK 7/8, 18 and 19 indicating a differentiated glandular phenotype, a finding associated with good prognosis, ER positivity and older patient age. In contrast, basal marker expression was significantly related to poor prognosis, ER negativity and younger patient age. Multivariate analysis showed that CK 5/6 was an independent indicator for relapse free interval. We were able to subgroup the cases into four distinct phenotype categories (pure luminal, mixed luminal/basal, pure basal and null), which had significant differences in relation to the biological features and the clinical course of the disease. Tumours classified as expressing a basal phenotype (the combined luminal plus basal and the pure basal) were in a poor prognostic subgroup, typically ER negative in most cases. These findings provide further evidence that breast cancer has distinct differentiation subclasses that have both biological and clinical relevance.