Cathepsin D as a potential prognostic marker for lung adenocarcinoma

We previously identified cathepsin D as a possible marker for lung adenocarcinoma (AD). The purpose of the present study is to evaluate the correlation between cathepsin D expression and clinicopathological findings or prognosis. We conducted immunohistochemistry (IHC) to assess 150 AD tissues. For these 150 tumors, TTF-1 expression, EGFR and KRAS gene mutations, and ALK rearrangements had already been examined. Cathepsin D expression was detected in 44% (66 of 150, IHC score ≥1+) and 27.3% (41 of 150, IHC score ≥2+). Cathepsin D-positive (IHC score ≥2+) tumors were more poorly differentiated than cathepsin D-negative ones, while all lepidic predominant invasive adenocarcinomas showed no cathepsin D expression. Univariate analysis revealed a poor prognosis for cathepsin D-positive lung AD patients with an IHC score ≥2+ (P=0.044). Cathepsin D expression was more frequent in TTF-1-negative than in TTF-1-positive ADs (P=0.034), and more frequent in ADs with EGFR wild genotype than mutant EGFR (P<0.001). Regarding AD patients with ALK rearrangements, 4 were positive for Cathepsin D, while 2 were negative. Cathepsin D expression is indicated to be a possible prognostic marker for lung AD and to correlate with a more poorly differentiated form.     

Immunohistochemical variability of epidermal growth factor receptor (EGFR) in liver metastases from colonic carcinomas

AIMS: To follow and compare immunohistochemical expression of epidermal growth factor receptor (EGFR) in tumour cells during the entire natural history of colonic carcinoma, from primary tumour to paired lymph node and sequentially resected liver metastases; and to test interobserver reproducibility of EGFR analysis. METHODS AND RESULTS: Forty patients had resection of colonic adenocarcinoma (27 with metastatic lymph nodes) and at least one partial hepatectomy (PH) for liver metastases; a second and a third PH were performed, respectively, in 14 and three patients; seven patients had tumour liver biopsy. EGFR immunohistochemistry (n = 130) was analysed independently by two pathologists. EGFR expression (membranous staining detected in > or = 1% of tumour cells) was detected in 38/40 colonic carcinomas, 23/26 lymph nodes and 51/64 liver metastases. Both primary tumours and related metastases were EGFR+ in 28 patients (73%). Discrepancies were found in EGFR status between liver and lymph node (23%) and among the different liver samples (31%). Interobserver agreement was very good (intraclass correlation coefficients of 0.81, 0.91 and 0.85, respectively, for interpretation of staining in colon, lymph node and liver metastases). CONCLUSIONS: Since immunohistochemical detection of EGFR remains a prerequisite for EGFR-targeted therapy eligibility, different tumour samples should be tested to allow every patient a chance to take advantage of this treatment.     

Change of HER-2/neu status in a subset of distant metastases from breast carcinomas

It is assumed that HER-2/neu status remains consistent in breast carcinoma during metastatic spread, but in most previous studies primary tumours were compared with concurrent regional lymph node metastases. The present study investigated 31 breast carcinomas and their corresponding lymph node and distant metastases for HER-2/neu status by immunohistochemistry (HercepTest) and fluorescence in situ hybridization (FISH) (PathVysion). In 14 cases, serum HER-2/neu (SHER-2) was measured sequentially using the Bayer Immuno 1 HER-2/neu assay. Comparing HER-2/neu immunohistochemistry of primary tumours and distant metastases case by case, increased HER-2/neu expression was found in the distant metastases in 15 cases (48.4%), three (9.7%) of which showed an increase from score 0 to score 3+. In contrast, lymph node metastases showed the same HER-2/neu expression as the primary tumours, confirmed by FISH. Two cases, which showed HER-2/neu score 3+ and HER-2/neu amplification in the primary tumours, revealed increased SHER-2 levels above 50 ng/ml at the first measurement. Five of the 14 cases (36%) showed an increase of SHER-2 above 50 ng/ml towards the end of the patients' life. On the basis of these results, there is evidence that in a subset of breast carcinomas, HER-2/neu amplification and overexpression occur de novo in distant metastases at a late disease stage.     

Study of molecular prognostic factors Bcl-2 and EGFR in rectal mucinous carcinomas

Colorectal cancer is an important public health problem worldwide, being the fourth most common cancer in men and the third most common in women. Colorectal cancer incidence is higher in developing countries due to the prevalence of obesity associated with reduced physical activity. Rectal mucinous carcinomas, especially the "signet ring cell" type, have a worse prognosis compared with other varieties of colorectal carcinomas, being diagnosed in more advanced stage and more prone to lymph node and peritoneal metastases. Our study comprised 37 cases with rectal adenocarcinoma with mucinous component operated in the Surgical Clinics of the Emergency County Hospital of Craiova, between 2006 and 2010. The aim of this study was to evaluate some molecular prognostic factors for rectal mucinous carcinomas namely B-cell lymphoma 2 (Bcl-2) and epidermal growth factor receptor (EGFR), and their correlations with the main morpho-clinical parameters of these patients. Thus, we immunohistochemically assessed the degree of apoptosis of tumor cells in mucinous rectal carcinomas using the Bcl-2 marker, and tumor aggressiveness using the EGFR responsiveness. In our study, the percentage of Bcl-2+ cases was 43.24%, with no significant statistical correlation between the Bcl-2 expression and histopathological subtype of mucinous adenocarcinoma. The evaluation of tumor aggressiveness in terms of EGFR responsiveness showed a reduced expression in carcinomas correlated with the increase in quantity of the mucinous component. In addition, EGFR reactivity was increased in the tumor invasion front.     

Remarkably high frequency of EGFR expression in breast carcinomas with squamous differentiation

The human epidermal growth factor receptor (EGFR) is reportedly overexpressed in 15-20% of breast carcinomas. EGFR overexpression is associated with reduced survival and is inversely correlated with expression of estrogen receptor (ER). This study assessed EGFR expression in breast carcinomas with squamous differentiation. The immunohistochemical (IHC) expression of EGFR was evaluated in 39 breast carcinomas with squamous differentiation (30 pure squamous, 6 adenosquamous, 3 carcinosarcomas) by use of the pharmDx assay (clone 2-18C9, DakoCytomation). Cases were considered positive if at least 10% of the cells showed 1+ positivity in the squamous component. Squamous differentiation was confirmed with IHC for CK5-6 (clone D5/16B4, DakoCytomation). ER, PR, and HER2 status as well as clinical information regarding treatment and outcome were correlated. As a control, a tissue microarray comprising 280 lymph node positive breast carcinomas was evaluated with the same EGFR assay. The 39 patients ranged in age from 33 to 77 years (mean 52). The tumors measured 1.3-30 cm (mean 4.8). Sentinel or full axillary lymph node dissection was performed in 28 patients. Fourteen patients had positive lymph nodes. At the time of initial diagnosis, 3 patients had distant metastasis. Follow-up was available for 16 patients (mean 45 months). Disease-free survival at 3 years was 70%. Among the 39 tumors 87% (34) were positive for EGFR (p<0.0001). Sixty-nine percent (27 of 39) showed >50% 2+ EGFR staining. EGFR-positive tumor cells (showing squamous morphology) were also found in 1 bone, 1 lung, and 8 of 11 lymph node metastases available for evaluation. All 11 lymph nodes showed squamous differentiation. All but 1 of the EGFR+ tumors examined were ER and PR negative. Six EGFR-positive tumors were HER2 positive. No statistically significant differences in HER2 status, size, lymph node status and disease-free survival were observed between EGFR+ and EGFR- cases, but the number of EGFR-negative tumors was quite small. Nine of 280 (3%) of lymph node-positive invasive carcinomas on the tissue microarray were EGFR+; review of the initial diagnostic slides failed to reveal squamous features in all but 1 of the 9 EGFR+ tumors. Breast carcinomas with squamous differentiation are a distinct subgroup of breast tumors with a very high frequency of EGFR positivity. Breast carcinomas of this type would be ideal candidates for a trial with EGFR inhibitors.     

Dipeptidyl peptidase IV expression in cancer and stromal cells of human esophageal squamous cell carcinomas,adenocarcinomas and squamous cell carcinoma cell lines

Dipeptidyl peptidase IV (DPPIV) is a transmembrane serine protease which is involved in the process of tumor invasion and development of metastases in human cancers. The aim of this study was to investigate the expression of DPPIV in cancer and stromal cells of both esophageal adenocarcinoma and squamous cell carcinoma (SCC). Tissue material from 159 patients was analyzed using immunohistochemistry. Western blotting was performed on cell lines and fresh frozen tissue sections. Results were compared with clinicopathological features. Evaluation of the immunohistochemical findings revealed significant differences between DPPIV expression in carcinoma cells and stromal cells, depending on the histological tumor type. A significantly higher level of DPPIV was found in adenocarcinomas compared to SCCs while no DPPIV was detected in normal esophageal epithelium. Overexpression of DPPIV in patients with adenocarcinoma was additionally associated with distant metastases. Thus, differences of DPPIV level in esophageal carcinomas compared with normal epithelium showed that esophageal malignancies were associated with an increased amount of cell surface‐bound DPPIV. Radiotherapy in patients had no impact on DPPIV expression in analyzed tissue samples. There was no correlation between DPPIV expression in cancer or stromal cells and survival of the patients.     

Expression of epidermal growth factor receptor in squamous cell carcinomas of the anal canal is independent of gene amplification.

Immunohistochemical detection of expression of the epidermal growth factor receptor (EGFR) has been utilized to identify eligible patients with solid malignant tumors, including colorectal adenocarcinoma, for monoclonal antibody therapy (eg, cetuximab). The EGFR status in squamous cell carcinoma of the anal canal, an uncommon malignancy traditionally treated with chemoradiation, has not been well investigated. In this study, 38 primary squamous cell carcinomas of the anal canal were immunohistochemically examined for EGFR expression and analyzed by fluorescence in situ hybridization (FISH) for EGFR gene copy numbers. The results showed a variable degree of EGFR expression in 21 (55%) tumors, among which 13 (62%) cases exhibited a 2+ to 3+ staining pattern according to the Dako EGFR phamDx interpretation guide. There were no significant differences among tumors stratified by stage, degree of keratinization, or tissue block storage times. FISH analysis showed that none of the 34 cases with interpretable results had EGFR gene amplification. Increased gene copy numbers due to polysomy 7 were seen in seven of 18 (39%) cases that expressed EGFR protein and four of 16 (25%) cases that did not (P=0.3876). Ten (56%) tumors with positive EGFR staining showed a balanced disomy 7 pattern and one case with monosomy 7 exhibited strong EGFR expression (3+). These results demonstrate that EGFR is overexpressed in more than one-half of the squamous cell carcinomas of the anal canal through mechanisms other than gene amplification. These observations may have important therapeutic implications since EGFR-based targeted therapies have shown promise for other malignant neoplasms.     

Nasopharyngeal carcinoma in children--a retrospective review and demonstration of Epstein-Barr viral genomes in tumor cell cytoplasm: a report of the Pediatric Oncology Group

Twenty-seven nasopharyngeal carcinomas were entered in the Pediatric Oncology Group Rare Tumor Registry from 1973 to 1988 (15 males, 12 females; 10 white, 15 black, two unknown; aged 8 to 17 years). Eight tumors were non-keratinizing carcinomas (World Health Organization 2) and 19 were undifferentiated (World Health Organization 3). The overall 3-year survival rate was 70% (SE 11%). Nine children developed distant metastases and two were salvaged. We found that localized tumor (P = .02) and black race (P = .05) were associated with a better outcome. In situ hybridization using a biotinylated probe demonstrated Epstein-Barr virus DNA in the cytoplasm of the neoplastic epithelial cells in nine of 11 tumors examined, firmly establishing the presence of Epstein-Barr virus within the malignant cells of nasopharyngeal carcinomas of both World Health Organization 2 and World Health Organization 3 histology, rather than in the surrounding lymphocytes.     

Overexpression of clusterin in human breast carcinoma

Clusterin has been implicated in numerous processes including active cell death, immune regulation, cell adhesion and morphological transformation. The purpose of this study was to examine clusterin expression in a large series of breast carcinomas by immunohistochemistry and in situ hybridization. The study included 40 samples of non-neoplastic glandular epithelia, 42 benign lesions, 15 atypical intraductal hyperplasias, 35 carcinomas in situ, 114 invasive carcinomas, and lymph node metastases from 40 patients. Epithelial normal cells were always negative for clusterin expression and only 19% of the benign lesions presented positive staining. In contrast to the benign lesions, however, the frequency of clusterin positive samples increased in atypical hyperplasias (47%, P = 0.08), intraductal carcinomas (49%, P = 0.01) and invasive carcinomas (53%, P < 0.001). Positive staining presented a cytoplasmic pattern, except in 3 cases of invasive carcinomas which had nuclear staining. Clusterin mRNA by in situ hybridization confirmed the specific cellular pattern of clusterin expression by immunohistochemistry. Clusterin expression was associated with large tumor size (P = 0.04), estrogen and progesterone receptor negative status (P = 0.02 and P = 0.001, respectively) and with the progression from primary carcinoma to metastatic carcinoma in lymph nodes (80% metastatic nodes had positive expression) (P = 0.004). Ten of 15 (67%) primary carcinomas without clusterin expression became positive in lymph node metastases, while most (22 of 25, 88%) of the clusterin-positive primary carcinomas were also immunoreactive in metastases. In survival analysis, clusterin expression did not represent a prognostic indicator by uni- or multivariate analysis. The increased clusterin expression in breast carcinomas tended to correlate inversely with the apoptotic index (P = 0.09) which indicates that clusterin gene expression is not a prerequisite to cellular death by apoptosis. From these results, we suggest that clusterin may have a role in tumorigenesis and progression of human breast carcinomas.     

Oxidative/nitrosative stress and peroxiredoxin 2 are associated with grade and prognosis of human renal carcinoma

Peroxiredoxins (Prxs) 1-6 were assessed in 138 renal cell carcinomas (RCC) using immunohistochemistry and selected samples by Western blotting analysis. Oxidative/nitrosative damage was evaluated using nitrotyrosine immunoreactivity. The expressions of Prxs were correlated with tumor grade and survival and nitrotyrosine reactivity. Non-malignant kidney tubular cells showed positivity with variable intensity for all six Prxs. In RCCs, most cases were positive for Prxs 1 and 2, while only 15-20% of tumors showed expression for Prxs 3 and 4. Prx 2 was associated with tumors of a lower grade (p=0.009) and with a lower frequency of distant metastases (p=0.046). Patients with tumors expressing Prx2 had better prognosis (p=0.027). Instead, nitrotyrosine was significantly associated with high grade tumors (p=0.001). Compared with the non-malignant kidney tubular cells, low Prx expression in the tumor cells can make them more susceptible to oxidative damage. Prx 2 was more abundantly expressed in low grade tumors, suggesting that this protein could play a role in preventing the development of oxidative damage, which in turn can lead to the activation of pathways leading to aggressive tumors.     

Prognostic value of immunocytochemical determination of parathyroid hormone-related peptide expression in cells of mammary ductal carcinoma. Analysis of 7 years of the disease course

Parathyroid hormone-related peptide (PTHrP) participates in the development of humoral hypercalcaemia of malignancy. The peptide is thought to affect growth and differentiation of normal and neoplastic cells. The present study aimed at evaluation of the relationship between survival time and development of distant metastases in patients with ductal mammary carcinoma on the one hand and PTHrP expression on the other. Immunocytochemical reactions using mouse monoclonal (clone 212-10.7) anti-PTHrP (38-64) antibodies were performed in paraffin sections originating from 47 patients with ductal mammary carcinoma. Expression of the protein was quantified employing a scale, considering the number of positive cells and intensity of the reaction (immunoreactive score, IRS). Survival time of the patients, determined during the course of a 7-year observation was also analysed. The obtained results demonstrated a relationship between intensity of PTHrP expression and the survival time. Patients with high expression of PTHrP (IRS>6) manifested longer survival than patients with lower PTHrP expression (IRS< or =6; Cox'es F test, P<0.05). Moreover, in the group with the lower PTHrP expression, a negative relationship was detected between expression of the protein and the survival time (Cox'es model, P<0.05). No relationship was detected between PTHrP expression and the development of distant metastases.     

Peripheral T Cell Lymphomas: An Immunological Study of Seven Unusual Cases

A multiparameter study of malignant lymph node cells and peripheral blood lymphocytes of seven patients with peripheral T cell lymphoma is presented. The results of monoclonal marker studies showed three cases of helper-suppressor T cell lymphoma (OKT4+, OKT8+), one case of suppressor T cell lymphoma (OKT8+), and three cases of helper T cell lymphoma (OKT4+). Immunophenotypic heterogeneity of neoplastic T cells with expression of pan-T antigens, OKT3+, and OKT11+ (erythrocyte rosetting+) was observed in most patients. Six of the seven cases tested showed Ia and DR antigens. No relationship was detected between patterns of reactivity with T cell reagents and histological types. When tested, the in-vitro malignant T cells of five patients proliferated in response to concanavalin A (Con A), but had poor response to phytohaemagglutinin. The interleukin 2 receptors showed maximum expression on Con A-activated T cells of five patients, and phytohaemagglutinin-activated T cells of one patient. The neoplastic T cells (OKT4+, OKT8+) of one patient studied had suppressor activity for IgG and IgA, and helper activity for IgM synthesis on pokeweed mitogen-induced normal B cell differentiations.     

P-glycoprotein expression in gastroesophageal adenocarcinomas, their metastases, and surrounding mucosa: a mapping study.

Previously, we identified P-glycoprotein in primary gastroesophageal adenocarcinomas and in adjacent mucosa. This study is a further investigation of P-glycoprotein expression in adenocarcinomas and benign mucosa. Sixteen resection specimens were studied (seven for gastric adenocarcinoma, seven for esophageal adenocarcinoma, one for adenocarcinoma at the gastroesophageal junction, and one for severe dysplasia in Barrett's esophagus). Multiple samples of tumor and mucosa were submitted according to a specimen diagram. Lymph node and distant metastases were studied when available. P-glycoprotein expression was identified in paraffin-embedded tissues by immunohistochemistry using monoclonal antibody C219 and was scored as the percentage of cells stained. P-glycoprotein was identified in six of 16 resection specimens. Intratumoral variability of C219 score was noted in three resections. No increase in expression was identified in lymph node or distant metastases as compared with primary tumors or in the invasive margin of the tumor as compared to the center. For every case in which tumors expressed P-glycoprotein, it was also diffusely present in all types of benign gastric and Barrett's mucosa, both adjacent to and distant from (up to 8 cm) the tumor. We also studied biopsies from 10 patients with Barrett's esophagus who did not have carcinoma. P-glycoprotein was only focally present in one of the 10 biopsies. Mucosa expressing P-glycoproteins may be the substrate from which a P-glycoprotein positive tumor arises.     

EGFR gene copy number increase in vulvar carcinomas is linked with poor clinical outcome

EGFR copy number increases have been frequently reported in cancer including vulvar carcinomas. Co-amplification of cancer genes plays an important role in the development of many tumour types. To better understand the effect of EGFR aberrations on vulvar cancer phenotype and patient prognosis, the authors analysed EGFR copy number changes using fluorescence in situ hybridisation and EGFR expression by immunohistochemistry in a tissue microarray containing 183 squamous cell carcinomas of vulva. Furthermore, the authors analysed the co-amplification frequency of EGFR with HER2, CCND1, MYC and PIK3CA, respectively. EGFR copy number increase was found in 39.3% of the tumours. Seventeen per cent of vulvar carcinomas showed EGFR high polysomy including 9% with amplification of the EGFR gene. Copy number gain of the EGFR locus was associated with non-basaloid phenotype (p=0.03), high-tumour stage (p<0.001), human papillomaviruse negativity of tumours (p=0.04) and the number of lymph node metastases (p=0.02). EGFR protein expression was statistically correlated to EGFR copy number increase (p<0.05). The observed co-amplification rate of EGFR with all four additionally examined oncogenes was much higher than statistically expected. There was a highly significant association between EGFR copy number increase and CCND1 amplifications (p<0.001) as well as the total number of gene amplifications (p=0.04). EGFR copy number gains were significantly related to unfavourable patient outcome in univariate analysis and multivariate Cox regression analysis. In conclusion, EGFR copy number increases are detectable in a substantial proportion of vulvar carcinomas with relationships to advanced tumour stages and the development of lymph node metastases. EGFR copy number aberrations are connected to other gene amplifications and probably define an human papillomaviruses-independent pathway in the development of vulvar carcinomas. These data support the potential utility of EGFR inhibitors as a therapeutic alternative in a subset of vulvar carcinomas.     

Elastosis in lung carcinoma: Immunohistochemical, ultrastructural and clinical studies

Elastosis is the pathological finding of focal deposits of elastic fibers in abnormal amounts within tissue. It is well described in the case of infiltrating carcinoma of the breast, but elastosis in lung carcinoma has not been previously documented in detail. We investigated the characteristics of elastosis in lung carcinoma with light and electron microscopies, and immunohistochemistry for alpha-1-antitrypsin. A total of 184 surgically resected primary lung carcinomas were studied. Elastosis was detected in adenocarcinomas (85/106), squamous cell carcinomas (11/60) and adenosquamous carcinomas (5/7), but not in small-cell carcinomas (n = 4) or large-cell carcinomas (n = 5). The degree of elastosis in each case was divided into one of five grades, graded as 3+ to 1-. The score of elastosis was significantly higher in adenocarcinoma than that in squamous-cell carcinoma (P < 0.01). In the cases of adenocarcinoma, the mean score of elastosis in the well-differentiated type (WD n = 43) was higher than that in the moderately differentiated (MD) (n = 39; P = 0.012) and poorly differentiated (PD) types (n = 24; P < 0.01). The mean score of elastosis in MD adenocarcinoma was also higher than that in the PD type (P < 0.01). Light- and electron-microscopic analyses revealed that these elastic fibers in elastosis were composed of aggregates of thick mature and fine immature elastic fibers, and were positive for alpha-1-antitrypsin. It is suggested that both degraded elastic fibers and newly synthesized fibers are contained in the elastosis of lung carcinoma. Although no significant evidence was detected to suggest any correlation between elastosis and the degree of tumor invasion, the survival curves of adenocarcinomas with elastosis showed a significantly improved prognosis than of those without elastosis in the cases of stages IA and IB (n = 52; P = 0.026).     

Osteonectin expression correlates with clinical outcome in thin cutaneous malignant melanomas

Osteonectin, also termed BM40 or SPARC (secreted protein, acidic and rich in cysteine) is a multifunctional glycoprotein involved in tissue mineralization, cell-extracellular matrix interactions as well as angiogenesis. It has been suggested that osteonectin may play a key role in the process of tumoral invasion and metastasis in certain malignancies. In this study, we reviewed the clinical records and the histopathologic slides of 188 thin cutaneous malignant melanomas (< or = 0.75 mm). Among them, 12 cases underwent progression and were selected for the study. Osteonectin expression was investigated by immunohistochemistry in these 12 patients and 24 matched controls who did not undergo progression. Osteonectin staining was correlated with clinical outcome and other clinicopathologic parameters. Progression-free and disease-specific survival rates were calculated with the Kaplan-Meier method and their differences were evaluated by the log rank test. Overall, immunoreactivity for osteonectin was found in 23 (63.8%) cases. Eighteen cases (50%) displayed staining in 1% to 50% of neoplastic cells whereas five cases (13.8%) showed a diffuse positivity in more than 50% of the tumor cells. Osteonectin expression was significantly correlated with risk of progression (P = .01), incidence of distant metastases (P = .005) and survival (P = .03). There was a higher incidence of osteonectin-positive tumors in cases that did experience regional lymph node metastases versus those cases that did not, but that difference did not reach statistical significance (P = .06). No significant correlation was found between osteonectin expression and other clinicopathologic features, including age, sex, site, histotype, Clark's level, presence of regression, presence of inflammatory response, and tumor growth phase. Our data showed that osteonectin expression is a predictor of clinical outcome in thin cutaneous melanomas.     

Clinical significance of CK5/6 and PTEN protein expression in patients with bilateral breast carcinoma

AIMS: To assess the expression of cytokeratin (CK) 5/6 in bilateral breast cancers and to assess the relationship between its expression and other prognostic variables, as well as between CK5/6 expression and patients' survival. METHODS AND RESULTS: The expression of CK5/6, PTEN protein, oestrogen receptor, progesterone receptor, p53 and c-erbB-2 protein were evaluated by immunohistochemistry in 88 primary breast cancers diagnosed in 44 women. To assess the prognostic value of studied factors, Cox regression analysis was performed. Expression of CK5/6 was found in 23 of 88 primary breast carcinomas (23/88; 26%). The hazard ratio of development of distant metastasis in patients in whom at least one cancer was CK5/6+ was 99.8 (P=0.037) and in patients with at least one carcinoma with reduced PTEN expression it was 10.8 (P=0.044). CK5/6 expression was correlated with absence of oestrogen (P<0.0001) and progesterone receptors (P<0.0001) and very strong expression of p53 (P<0.05). Reduced PTEN expression was correlated with presence of axillary metastases (P<0.01), with very strong expression of c-erbB-2 (P<0.05) and with reduced expression of oestrogen receptor (P<0.05). CONCLUSIONS: Analysis of expression of CK5/6 and PTEN protein in bilateral breast carcinomas may be of value in clinical practice and warrant further studies.     

Overexpression of claudin proteins in esophageal adenocarcinoma and its precursor lesions.

Claudins are components of tight junctions important in intercellular barriers and cell polarity. The authors identified upregulation of Claudins 3, 4, and 7 in gastric adenocarcinoma using Affymetrix U-133 oligonucleotide microarrays and immunohistochemistry (IHC). While normal gastric mucosa lacked Claudin 3, 4, and 7 expression, intestinal metaplasia and dysplasia showed these proteins. The authors hypothesized that Claudins would be similarly overexpressed in Barrett's esophagus (BE)/adenocarcinoma. Claudins 3, 4, and 7 gene expression was analyzed by Affymetrix U-133 microarrays in three esophageal adenocarcinomas, one case of BE, and three normal esophagi. IHC validation was performed using tissue microarrays constructed from esophageal resection specimens containing squamous (44 cases), gastric (40 cases), and non-dysplastic BE (16 cases), low-grade and high-grade dysplasia (16 and 26 cases), adenocarcinoma (58 cases), and nodal metastases (27 cases). IHC staining was scored semiquantitatively (0+ to 4+). By microarray analysis, Claudin 3 showed a marked increase in mRNA expression compared with normal esophagus (approximately 100-fold). Claudins 4 and 7 were modestly increased (2.2- and 1.3-fold). By IHC, Claudin 3 expression was 1+ in most (>95%) normal squamous or gastric tissues and 2+ to 4+ in more than 80% of high-grade dysplasia, adenocarcinoma, and metastases specimens. Claudin 4 protein expression was 2+ or less in most squamous and gastric mucosa (>90%) but 3+ or 4+ in BE, low- and high-grade dysplasia, adenocarcinoma, and metastases specimens (>90%). Claudin 7 expression was minimal in squamous and gastric mucosa but strong (3+ to 4+) in BE and low-grade dysplasia. In high-grade dysplasia, adenocarcinoma, and metastases, Claudin 7 was less intense, with 60% to 70% staining 3+ or 4+ and 30% to 40% staining weakly (1+ or 2+). The findings suggest that alterations in Claudin proteins are an early event in tumorigenesis and may provide targets for diagnosis and directed therapy for esophageal adenocarcinoma and its precursors.