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β-Catenin Expression Pattern in Stage I and II Ovarian Carcinomas : Relationship with β-Catenin Gene Mutations, Clinicopathological Features, and Clinical Outcome

The immunohistochemical expression pattern of β-catenin has been correlated with β-catenin gene mutations, clinicopathological features, and disease outcome in 69 stage I and II ovarian carcinomas. β-Catenin expression was localized in the nuclei, in addition to the cytoplasm and membrane, in 11 tumors (16%). nine endometrioid carcinomas with widespread nuclear expression and two serous carcinomas with focal nuclear expression. The remaining 58 carcinomas (84%) only had membranous β-catenin expression. All but one of the endometrioid carcinomas with nuclear β-catenin expression had considerable squamous metaplasia, and five of these cases had large areas of endometrioid tumor of low malignant potential. In addition, β-catenin nuclear expression was observed in atypical epithelial cells in endometriotic glands adjacent to an endometrioid carcinoma. Sequencing was performed on 25 tumors and corresponding normal tissue: all 13 endometrioid tumors as well as 12 carcinomas of other histological types (four serous, two clear cell, two mucinous, and two mixed). There were oncogenic mutations in the phosphorylation sequence for GSK-3β in exon 3 of the β-catenin gene in seven endometrioid carcinomas with β-catenin nuclear expression. Three mutations affected codon 32 (D32G, D32Y, and D32Y), one affected codon 33 (S33C), two affected codon 37 (S37C and S37F), and one affected codon 41 (T41A). No mutations were observed in the other 18 carcinomas analyzed, comprising two endometrioid and two serous carcinomas with β-catenin nuclear expression, and 14 carcinomas of different histological types with only membranous expression. In the univariate and multivariate survival analyses, β-catenin nuclear expression was selected as an indicator of good prognosis, because no patient whose tumor expressed β-catenin in the nuclei showed relapses or died, in contrast to the 19 relapses and deaths among patients with tumors that only had β-catenin membranous expression, including three of the four patients with endometrioid carcinomas. Oncogenic β-catenin mutation is characteristic of a group of endometrioid carcinomas with a good prognosis, most of which originate from previous benign or borderline lesions. Endometrioid carcinomas with exclusively membranous expression of β-catenin seem to represent a different subgroup of carcinomas that probably have a worse prognosis. In early-stage ovarian cancer, determination of the β-catenin expression pattern could prove to be a useful marker for selecting low-risk patients.

Cytoplasmic localization of p120ctn and E-cadherin loss characterize lobular breast carcinoma from preinvasive to metastatic lesions

Accumulating evidences indicate that p120 catenin, a member of the E-cadherin (E-CD)/catenin adhesion complex, plays a role in tumor invasion. To establish the expression pattern of p120 in breast cancer, we analysed 326 breast tissue biopsies by tissue microarray. Most of the lobular tumors (88%) showed exclusive cytoplasmic localization, and 6% of them also had p120 nuclear staining. Cytoplasmic p120 strongly associated with complete loss of E-CD and β-catenin not only in lobular carcinoma and its metastases but also in atypical lobular hyperplasias. In the latter, loss of heterozygosity of E-CD gene was also observed. Complete loss of E-CD and cytoplasmic and nuclear p120 staining was also observed in primary lobular cancer cell cultures generated by us. In ductal tumors, by contrast, reduction of p120 and E-CD in membrane was very common (57 and 53%, respectively), whereas cytoplasmic p120 staining was rarely seen. This simultaneous reduction of membranous E-CD and p120 was not associated with increased Src kinase activity. To demonstrate that cytoplasmic p120 localization was a consequence of the absence of E-CD, the endogenous E-CD was re-expressed in MDA-231 cells by 5-Aza-2′-deoxycytidine (5Aza) treatment. After treatment, p120 shifted from the cytoplasm to the membrane, where it colocalized with endogenous E-CD. Additionally, suppressing E-CD expression in Madin–Darby canine kidney cells by stable transfection of the transcriptional repressors Snail, E47 or Slug, provokes p120 cytoplasmic localization and p120 isoform switching. In conclusion, abnormal cytoplasmic and nuclear localization of p120, which are mediated by the absence of E-CD, characteristically occur in the early stages of lobular breast cancer and are maintained during tumor progression to metastasis. Consequently, p120 may be an important mediator of the oncogenic effects derived from E-CD inactivation, including enhanced motility and invasion, in lobular breast cancer.

β-catenin Expression Pattern, β-catenin Gene Mutations, and Microsatellite Instability in Endometrioid Ovarian Carcinomas and Synchronous Endometrial Carcinomas

&bgr;-Catenin gene mutations and microsatellite instability (MI) have been reported in endometrioid ovarian carcinomas. In colon but not endometrial cancer, &bgr;-catenin gene mutations are associated with a replication error phenotype and MI. In this study the authors investigate whether &bgr;-catenin mutations and MI are two independent oncogenic pathways in endometrioid ovarian carcinomas. They also evaluate the usefulness of these molecular markers in determining the primary origin of simultaneous tumors in the ovary and endometrium. This study was performed on 26 patients diagnosed with primary endometrioid ovarian carcinoma, five of whom also had pathologically diagnosed primary synchronous endometrioid endometrial carcinoma. Immunohistochemical and molecular analyses indicated that there were 25 primary ovarian tumors with four primary synchronous endometrial cancers and one ovarian metastasis of a primary endometrial carcinoma. All studies were performed on formalin-fixed, paraffin-embedded tissue samples. The &bgr;-catenin expression pattern (nuclear vs. membranous) was analyzed immunohistochemically. Mutations in exon 3 of the &bgr;-catenin gene were studied by polymerase chain reaction, single-strand conformational polymorphism, and direct sequencing. MI status was established by studying BAT-26 and BAT-25 mononucleotide repeats. In the group with 21 single ovarian tumors, 18 (85%) had &bgr;-catenin nuclear expression, eight (38%) had &bgr;-catenin gene mutations (always associated with &bgr;-catenin nuclear expression), and four (19%) had MI. Only one case (5%) had both &bgr;-catenin gene mutations and MI. The mutations affected one of the serine/threonine residues targeted for phosphorylation by glycogen synthase kinase-3&bgr; or adjacent residues. At codon 32, a GAC-to-TAC (D32Y) change was found; at codon 33, two TCT-to-TGT (S33C) changes were found; at codon 37, three TCT-to-TTT (S37F) changes and one TCT-to-TGT (S37C) change were found; and, lastly, one ACC-to-GCC change at codon 41 (T41A) was detected. Four of the 25 endometrioid ovarian carcinomas (16%) had an associated synchronous endometrial carcinoma. There was a higher percentage of &bgr;-catenin mutations (n = 3, 75%) in synchronous ovarian carcinomas than in single ones, although with a similar percentage of MI (n = 1, 25%). &bgr;-catenin mutations were S37C in two cases and D32G in one. One of the four endometrial carcinomas showed an S33C &bgr;-catenin mutation, and two carcinomas had MI. None of the four tumors had both &bgr;-catenin gene mutation and MI. &bgr;-catenin gene mutations were always associated with a nuclear &bgr;-catenin expression pattern, whereas MI was associated with a membranous pattern. In one patient both the ovarian and the endometrial carcinomas had &bgr;-catenin gene mutations, in another patient both tumors showed MI, whereas in the remaining two patients the ovarian carcinomas showed &bgr;-catenin gene mutations and the endometrial carcinomas showed MI. To summarize, the results of this study suggest that &bgr;-catenin mutations and MI could represent two independent pathways in endometrioid ovarian carcinomas because they occur simultaneously very infrequently (in 5% of these cases). &bgr;-catenin mutations are always associated with a nuclear &bgr;-catenin expression pattern, whereas cases with a replication error -plus phenotype showed no abnormal &bgr;-catenin subcellular localization. The study of the &bgr;-catenin expression pattern, &bgr;-catenin mutations, and MI, together with conventional clinicopathologic findings, could aid in distinguishing between the metastatic or independent origin of simultaneous endometrioid ovarian and endometrial carcinomas. Tumors with identical immunohistochemical and molecular features should therefore be considered to have a common origin.

Breast Cancer Prognosis Determined by Gene Expression Profiling: A Quantitative Reverse Transcriptase Polymerase Chain Reaction Study

PURPOSE We sought to reproduce with quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) the results obtained with a 70-gene expression profile that has been described previously in breast cancer. PATIENTS AND METHODS Frozen breast cancer samples from patients who were operated on were used to isolate tumor RNA. Ninety-six patients with stage I to II disease were included. Median age was 57 years (range, 27 to 80 years). Forty-eight patients had lymph node-negative and 48 lymph node-positive disease. qRT-PCR amplifications were performed and the results were correlated with clinical data. RESULTS After a minimum follow-up of 5 years, 25 patients had a relapse. The gene profile divided patients into two groups with poor and good prognosis. Significant differences with regard to grade of differentiation, size and hormone receptors were seen between the two groups. The gene profile was significantly associated with relapse-free survival and overall survival in the whole group of 96 patients. Multivariate analysis showed that only lymph node status and gene profile were significantly correlated to overall survival. CONCLUSION qRT-PCR reproduced the results obtained with microarrays for a prognostic gene profile in women with early-stage breast cancer.

Aurora kinases as prognostic biomarkers in ovarian carcinoma.

We investigated the expression of Aurora kinases A and B by immunohistochemistry in 68 ovarian carcinomas to analyze their prognostic value. The amplification of AURKA gene by fluorescence in situ hybridization was also assessed. Overall, 58.8% and 85.3% of ovarian carcinomas showed expression of Aurora A and B, respectively. Amplification of AURKA was found in 27.6% of cases examined. Tumors with Aurora A expression showed a lower rate of recurrence than those tumors without Aurora A expression (65% versus 91.7%, P = .019). In the univariate analysis, patients with Aurora A and B expression showed an increased progression-free survival (P = .023 and .06, respectively, log-rank test) and overall survival (P = .03 and .02, respectively, log-rank test). The multivariate analysis adjusted to optimal surgery by Cox proportional hazards regression showed Aurora A expression as an independent prognostic factor for progression-free survival (P = .03) and overall survival (P = .02). In conclusion, Aurora A expression seems to have a prognostic value in ovarian carcinoma.

Signet-ring stromal tumor of the ovary: A histochemical, immunohistochemical and ultrastructural study

We report a signet-ring stromal tumour found in the ovary of a 50-year-old woman. The solid tumour was composed of spindle and round cells which stained with vimentin and some round cells were vacuolated and had the appearance of signet-ring cells. The vacuoles were negative with periodic acid-Schiff, with and without diastase digestion, with alcian blue, Sudan III, alpha-1-antitrypsin and alpha-fetoprotein stains. Ultrastructurally, they were membrane bound and contained scanty flocculent material. Some neoplastic cells contained intracytoplasmic eosinophilic globules which were positive with alpha-1-antitrypsin. Signet-ring stromal tumour is a benign, morphologically distinctive type of ovarian stromal tumour, which may present problems in differential diagnosis with other stromal and non stromal ovarian neoplasms.

Relationship between ERBB2 and E-cadherin expression in human breast cancer.

A recent in vitro study has suggested that overexpression of ERBB2 may mediate breast tumour progression and metastasis by inhibiting the transcription of the E-cadherin (E-CD) gene. To test this hypothesis in human breast cancer in vivo, we studied the relationship between the expression of both molecules in 247 breast carcinomas immunohistochemically. Five ductal carcinomas in situ overexpressed ERBB2 and showed preserved E-CD expression. Forty-four of 226 infiltrating ductal carcinomas (19.47%) showed ERBB2 overexpression, and a statistically significant relationship was found between ERBB2 overexpression and high histological grade. E-CD expression was preserved in 111 cases (49.1%) and correlated with the histological grade. However, no significant relationship was found between ERBB2 and E-CD expression. None of the 16 infiltrating lobular carcinomas expressed ERBB2 or E-CD. These observations in different histological types of breast carcinoma strongly argue against a role for ERBB2 as a transcriptional regulator of E-CD expression in most human breast carcinomas in vivo.

Angiogenesis-related gene expression profile with independent prognostic value in advanced ovarian carcinoma.

Background Ovarian carcinoma is the most important cause of gynecological cancer-related mortality in Western societies. Despite the improved median overall survival in patients receiving chemotherapy regimens such as paclitaxel and carboplatin combination, relapse still occurs in most advanced diseased patients. Increased angiogenesis is associated with rapid recurrence and decreased survival in ovarian cancer. This study was planned to identify an angiogenesis-related gene expression profile with prognostic value in advanced ovarian carcinoma patients. Methodology/Principal Findings RNAs were collected from formalin-fixed paraffin-embedded samples of 61 patients with III/IV FIGO stage ovarian cancer who underwent surgical cytoreduction and received a carboplatin plus paclitaxel regimen. Expression levels of 82 angiogenesis related genes were measured by quantitative real-time polymerase chain reaction using TaqMan low-density arrays. A 34-gene-profile which was able to predict the overall survival of ovarian carcinoma patients was identified. After a leave-one-out cross validation, the profile distinguished two groups of patients with different outcomes. Median overall survival and progression-free survival for the high risk group was 28.3 and 15.0 months, respectively, and was not reached by patients in the low risk group at the end of follow-up. Moreover, the profile maintained an independent prognostic value in the multivariate analysis. The hazard ratio for death was 2.3 (95% CI, 1.5 to 3.2; p<0.001). Conclusions/Significance It is possible to generate a prognostic model for advanced ovarian carcinoma based on angiogenesis-related genes using formalin-fixed paraffin-embedded samples. The present results are consistent with the increasing weight of angiogenesis genes in the prognosis of ovarian carcinoma.