Nidogen 1 and Nuclear Protein 1: novel targets of ETV5 transcription factor involved in endometrial cancer invasion.

Endometrial cancer is the most frequent malignancy of the female genital tract in western countries. Our group has previously characterized the upregulation of the transcription factor ETV5 in endometrial cancer with a specific and significant increase in those tumor stages associated with myometrial invasion. We have shown that ETV5 overexpression in Hec1A endometrial cancer cells induces epithelial to mesenchymal transition resulting in the acquisition of migratory and invasive capabilities. In the present work, we have identified Nidogen 1 (NID1) and Nuclear Protein 1 (NUPR1) as direct transcriptional targets of ETV5 in endometrial cancer cells. Inhibition of NID1 and NUPR1 in ETV5 overexpressing cells reduced cell migration and invasion in vitro and reduced tumor growth and dissemination in an orthotopic endometrial cancer model. Importantly, we confirmed a significant increase of NUPR1 and NID1 protein expression in the invasion front of the tumor compared to their paired superficial zone, concomitant to ETV5 overexpression. Altogether, we conclude that NID1 and NUPR1 are novel targets of ETV5 and are actively cooperating with ETV5 at the invasion front of the tumor in the acquisition of an invasive phenotype to jointly drive endometrial cancer invasion.

Stromal signatures in endometrioid endometrial carcinomas.

The pattern of myometrial invasion in endometrioid endometrial carcinomas varies considerably; ie, from widely scattered glands and cell nests, often associated with a fibromyxoid stromal reaction (desmoplasia) and/or a lymphocytic infiltrate, to invasive glands with little or no stromal response. Recently, two distinct stromal signatures derived from a macrophage response (colony-stimulating factor 1, CSF1) and a fibroblastic response (desmoid-type fibromatosis, DTF) were identified in breast carcinomas and correlated with clinicopathologic features including outcome. In this study, we explored whether these stromal signatures also apply to endometrioid carcinomas and how their expression patterns correlated with morphologic changes. We studied the stromal signatures both by immunohistochemistry and in situ hybridization in 98 primary endometrioid carcinomas with (87 cases) and without (11 cases) myometrial invasion as well as in the corresponding regional lymph nodes metatases of 9 myoinvasive tumors. Desmoplasia correlated positively with the DTF expression signature. Likewise, mononuclear infiltrates were found in the stroma of tumors expressing CSF1. Twenty-four out of eighty-seven (27%) myoinvasive endometrioid carcinomas were positive for the macrophage signature and thirteen out of eighty-seven (15%) expressed the fibroblast signature. Eleven additional cases were positive for both DTF and CSF1 signatures (11/87; 13%). However, over half of the cases (39/87; 45%) and the majority of the non-myoinvasive tumors (8/11; 73%) failed to express any of the two stromal signatures. The macrophage response (CSF1) was associated with higher tumor grade, lymphovascular invasion, and PIK3CA mutations (P<0.05). There was a concordance in the expression of the CSF1 signature in the primary tumors and their corresponding lymph node metastases. This study is the first characterization of stromal signatures in endometrioid carcinomas. Our findings shed new light on the relationship between genetically different endometrioid carcinomas and various stromal responses. Preservation of the CSF1 macrophage stromal response in the metastases leds support to targeting the CSF1 pathway in endometrioid endometrial carcinomas.

Cyclooxygenase-2 inhibitor suppresses tumour progression of prostate cancer bone metastases in nude mice.

OBJECTIVE: To assess whether celecoxib, a selective cyclooxygenase-2 (COX-2) inhibitor with anti-cancer properties, has an inhibitory effect on tumour establishment and progression of prostate cancer (PCa) bone metastases. MATERIALS AND METHODS: PC-3 stable luciferase-expressing cells were injected into male nude mice by intracardiac (i.c.) and intratibial (i.t.) injections, and the effect of celecoxib on bone metastases was then recorded using bioluminescence image analysis. In cases of chemoprevention, mice received 3 mg/kg celecoxib from 1 week before cell implantation until the end of the study, and to test the therapeutic effect, mice received celecoxib 1 week after cell implantation until the end of the study. Tumour tissue samples were histologically examined and COX-2 expression was quantified at the protein level. RESULTS: Celecoxib significantly decreased cell viability and the proliferation of human PCa cells in vitro in a dose-dependent manner. Bone metastases were detected after i.c. injection in nude mice. Celecoxib (15 ppm) administered before i.c. injection did not inhibit the cellular metastatic potential, as the numbers of bone metastases were similar in both groups. However, celecoxib did decrease metastatic progression in the osseous environment when cells were injected directly into the tibia (P < 0.05). At the protein level, COX-2 expression was significantly decreased in the celecoxib treatment group (P < 0.01). CONCLUSION: In a preclinical mice model, celecoxib administered orally at the standard human dose inhibits the progression of established PCa bone metastases.

Molecular diagnosis of endometrial cancer from uterine aspirates.

Rapid and reliable diagnosis of endometrial cancer (EC) in uterine aspirates is highly desirable. Current sensitivity and failure rate of histological diagnosis limit the success of this method and subsequent hysteroscopy is often necessary. Using quantitative reverse transcriptase-polymerase chain reaction on RNA from uterine aspirates samples, we measured the expression level of 20 previously identified genes involved in EC pathology, created five algorithms based on combinations of five genes and evaluated their ability to diagnose EC. The algorithms were tested in a prospective, double-blind, multicenter study. We enlisted 514 patients who presented with abnormal uterine bleeding. EC was diagnosed in 60 of the 514 patients (12%). Molecular analysis was performed on the remnants of aspirates and results were compared to the final histological diagnoses obtained through biopsies acquired by aspiration or guided by hysteroscopy, or from the specimens resected by hysterectomy. Algorithm 5 was the best performing molecular diagnostic classifier in the case-control and validation study. The molecular test had a sensitivity of 81%, specificity of 96%, positive predictive value (PPV) of 75% and negative predictive value (NPV) of 97%. A combination of the molecular and histological diagnosis had a sensitivity of 91%, specificity of 97%, PPV of 79% and NPV of 99% and the cases that could be diagnosed on uterine aspirate rose from 76 to 93% when combined with the molecular test. Incorporation of the molecular diagnosis increases the reliability of a negative diagnosis, reduces the need for hysteroscopies and helps to identify additional cases.

The EMT signaling pathways in endometrial carcinoma.

Endometrial cancer (EC) is the most common gynecologic malignancy of the female genital tract and the fourth most common neoplasia in women. In EC, myometrial invasion is considered one of the most important prognostic factors. For this process to occur, epithelial tumor cells need to undergo an epithelial to mesenchymal transition (EMT), either transiently or stably, and to differing degrees. This process has been extensively described in other types of cancer but has been poorly studied in EC. In this review, several features of EMT and the main molecular pathways responsible for triggering this process are investigated in relation to EC. The most common hallmarks of EMT have been found in EC, either at the level of E-cadherin loss or at the induction of its repressors, as well as other molecular alterations consistent with the mesenchymal phenotype-like L1CAM and BMI-1 up-regulation. Pathways including progesterone receptor, TGFß, ETV5 and microRNAs are deeply related to the EMT process in EC.

Molecular bases of endometrial cancer: new roles for new actors in the diagnosis and the therapy of the disease.

Endometrial carcinoma (EC) is the most commonly diagnosed gynecologic malignancy in the western world. The majority of these cancers are curable, but a subset about 15-20% of endometrial tumors exhibits an aggressive phenotype. Based on clinic-pathological and molecular characteristics, EC has been classified into two groups: Type I estrogen-dependent adenocarcinomas, which have a good prognosis and an endometrioid histology, and Type II or non-estrogen-dependent EC associated with poor prognosis and non-endometrioid histology. EC develops as a result of a stepwise accumulation of alterations that seem to be specific of each histological type. However, more knowledge is needed to better understand the differences in the biology and the clinical outcome of EC. We would like to highlight the need to explore new potential biomarkers of EC as a tool for the detection and monitoring of aggressive endometrial tumors that, at the same time, will allow us to develop novel and more selective molecular targeted therapies against EC.

A three-gene panel on urine increases PSA specificity in the detection of prostate cancer.

BACKGROUND: Several studies have demonstrated the usefulness of monitoring an RNA transcript, such as PCA3, in post-prostate massage (PM) urine for increasing the specificity of prostate-specific antigen (PSA) in the detection of prostate cancer (PCa). However, a single marker may not necessarily reflect the multifactorial nature of PCa. METHODS: We analyzed post-PM urine samples from 154 consecutive patients, who presented for prostate biopsies because of elevated serum PSA (>4 ng/ml) and/or abnormal digital rectal exam. We tested whether the putative PCa biomarkers PSMA, PSGR, and PCA3 could be detected by quantitative real-time PCR in post-PM urine sediment. We combined these findings to test if a combination of these biomarkers could improve the specificity of actual diagnosis. Afterwards, we specifically tested our model for clinical usefulness in the PSA diagnostic "gray zone" (4-10 ng/ml) on a target subset of 82 men with no prior biopsy. RESULTS: By univariate analysis, we found that the PSMA, PSGR, and PCA3 scores were significant predictors of PCa. Using a multiplex model, the area under the multi receiver-operating characteristic curve was 0.74 versus 0.82 in the diagnostic "gray zone." Fixing the sensitivity at 96%, we obtained a specificity of 34% and 50% in the gray zone. CONCLUSIONS: Taken together, these results provide a strategy for the development of a more accurate model for PCa diagnosis. In the future, a multiplexed, urine-based diagnostic test for PCa with a higher specificity, but the same sensitivity as the serum-PSA test, could be used to determine better which patients should undergo biopsy.

Molecular markers of endometrial carcinoma detected in uterine aspirates.

Endometrial cancer (EC) is the most frequent of the invasive tumors of the female genital tract. Although usually detected in its initial stages, a 20% of the patients present with advanced disease. To date, no characterized molecular marker has been validated for the diagnosis of EC. In addition, new methods for prognosis and classification of EC are needed to combat this deadly disease. We thus aimed to identify new molecular markers of EC and to evaluate their validity on endometrial aspirates. Gene expression screening on 52 carcinoma samples and series of real-time quantitative PCR validation on 19 paired carcinomas and normal tissue samples and on 50 carcinoma and noncarcinoma uterine aspirates were performed to identify and validate potential biomarkers of EC. Candidate markers were further confirmed at the protein level by immunohistochemistry and Western blot. We identified ACAA1, AP1M2, CGN, DDR1, EPS8L2, FASTKD1, GMIP, IKBKE, P2RX4, P4HB, PHKG2, PPFIBP2, PPP1R16A, RASSF7, RNF183, SIRT6, TJP3, EFEMP2, SOCS2 and DCN as differentially expressed in ECs. Furthermore, the differential expression of these biomarkers in primary endometrial tumors is correlated to their expression level in corresponding uterine fluid samples. Finally, these biomarkers significantly identified EC with area under the receiver-operating-characteristic values ranging from 0.74 to 0.95 in uterine aspirates. Interestingly, analogous values were found among initial stages. We present the discovery of molecular biomarkers of EC and describe their utility in uterine aspirates. These findings represent the basis for the development of a highly sensitive and specific minimally invasive method for screening ECs.

Subtractive proteomic approach to the endometrial carcinoma invasion front.

Tumor invasion defines the transition between tissue-restricted carcinomas, related to good outcome as optimal surgery becomes possible, and metastatic tumors associated with poor prognosis and a dramatic decrease in survival. In endometrial cancer, myometrial infiltration represents a determinant parameter highly valuable in prognosis. To date, the identification of proteins involved in endometrial carcinoma invasion has been essentially conducted by immunohistochemical methods, without a global perception on the invasive front. Laser microdissection presents nowadays limitations to the profound spatiotemporal regulation from both the tumor and the surrounding stroma occurring at the invasive front. In this work, we attempted an alternative proteomic approach to characterize specific components of the tumor invasive front or its reactive stroma, by comparing the invasive area of an endometrial carcinoma with the noninvasive superficial tumor area and normal tissue from the same patients. This strategy led us to identify proteins involved in cellular morphology, assembly and movement, differentially expressed at the invasive front, as well as pathways like cell-to-cell signaling and interaction and a modulated response to oxidative stress as events related to endometrial carcinoma invasion. In conclusion, we could identify new players of myometrial infiltration by applying a subtractive proteomic approach to the endometrial carcinoma invasion front.