[HE4, a novel marker for epithelial ovarian cancer: evaluation of analytical performances]. / HE4, un nouveau marqueur des cancers épithéliaux ovariens: évaluation des performances analytiques.

Human epididymis protein 4 (HE4) is a novel marker for ovarian cancer. HE4 exhibits a high sensitivity to detect ovarian cancer and can be used with CA125 as a predictor of malignancy. Additional uses of HE4 are as an aid of monitoring response to therapy for patients with invasive ovarian cancer and as a marker to detect recurrences in the follow-up after treatment of the primary tumor. The HE4 EIA, an enzyme immunoassay for the quantitative determination of HE4 in human serum developed by Fujirebio Diagnostic Inc. (Tokyo, Japan), is now available with a CE-IVD label in Europe (Diasource, Nivelles, Belgium). The aim of the study was to evaluate according to the COFRAC LAB GTA 04 guide, the analytical performance of the test, using 4 standardized samples (target values: 49.8, 140.4, 167.6 and 415.2 pmol/L) and serum samples from patients with ovarian cancer treated in our institution. Intra- and inter-assay precisions showed coefficients of variation less than 10%. The low limit of detection (4 pmol/L) and the limit of quantitation (8 pmol/L) are suitable for clinical samples assessment. The assay mean dilution linearity is 100 +/- 10% (90 to 107% of recovery). Globally, the uncertainty varied from 13.1% (low values) to 28.1% (elevated values). We conclude that the HE4 EIA from Fujirebio Diagnostic Inc. displayed convenient analytical performances that allows its use it clinical practice.

Oestrogen receptor negative breast cancers exhibit high cytokine content.

INTRODUCTION: An emerging hypothesis suggests that cytokines could play an important role in cancer as potential modulators of angiogenesis and leucocyte infiltration. METHODS: A novel multiplexed flow cytometry technology was used to measure the expression of 17 cytokines (IL-1beta, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10, IL-12 [p70], IL-13, IL-17, granulocyte colony-stimulating factor [CSF], granulocyte-macrophage CSF, IFN-gamma, monocyte chemoattractant protein [MCP]-1, macrophage inflammatory protein [MIP]-1beta, tumour necrosis factor [TNF]-alpha) at the protein level in 105 breast carcinoma. B lymphocyte, T lymphocyte and macrophage levels were determined by immunohistochemistry. RESULTS: Fourteen of the 17 cytokines were expressed in breast carcinoma, whereas only nine cytokines could be detected in normal breast. Most cytokines were more abundant in breast carcinoma than in normal breast, with IL-6, IL-8, granulocyte CSF, IFN-gamma, MCP-1 and MIP-1beta being very abundant. IL-2, IL-6, IL-8, IL-10, IFN-gamma, MCP-1, MIP-1beta and TNF-alpha, and to a lesser extent IL-1beta and IL-13 exhibited levels of expression that were inversely correlated to oestrogen receptor and progesterone receptor status. Most cytokines were not correlated with age at cancer diagnosis, tumour size, histological type, or lymph node status. However, IL-1beta, IL-6, IL-8, IL-10, IL-12, MCP-1 and MIP-1beta were more abundant in high-grade tumours than in low-grade tumours. In addition, IL-8 and MIP-1beta were expressed to a greater degree in HER2-positive than in HER2-negative patients. The expression of most of the studied cytokines was correlated to levels of activator protein-1, which is known to regulate numerous cytokines. Overexpression of MCP-1 and MIP-1beta were linked to B lymphocyte, T lymphocyte and macrophage infiltration, whereas high levels of IL-8 were correlated with high macrophage content in tumour. Moreover, IL-8 positive tumours exhibited increased vascularization. CONCLUSION: We found that multiple cytokines were overexpressed in oestrogen receptor negative breast carcinoma, and that the three major cytokines--MCP-1, MIP-1beta and IL-8--were correlated with inflammatory cell component, which could account for the aggressiveness of these tumours.

Expression and biological role of the prostaglandin D synthase/SOX9 pathway in human ovarian cancer cells.

New therapeutic strategies for ovarian cancer include the identification of involved signaling pathways that could potentially serve as a source of biomarkers for early stages of the disease. In this study, we show that the embryonic male prostaglandin D synthase (Pgds)/SOX9 pathway is expressed at both the RNA and protein levels in different types of human ovarian tumors, pointing to Pgds and SOX9 as possible diagnostic markers for ovarian carcinomas. Using ovarian cancer cell lines, we found, first, that components of the Pgds/SOX9 pathway are expressed in these cells, and second, that treatment of these cells with prostaglandin D2 (PGD2) can inhibit their growth via its DP1 receptor and induce apoptosis. Finally, using siRNA and overexpression strategies, we demonstrate that SOX9 expression is induced by PDG2 and is responsible for PDG2-mediated growth inhibition. Accordingly, as stimulating the PGD2/DP1 signal transduction pathway upregulates SOX9 expression, either activators of this pathway or DP1 agonists may be useful as new therapeutic agents.

The anti-Müllerian hormone type II receptor: insights into the binding domains recognized by a monoclonal antibody and the natural ligand.

Anti-Müllerian hormone (AMH) [also called Müllerian inhibiting substance (MIS)] is a member of the transforming growth factor-beta family. AMH and its type II receptor (AMHR-II) are involved in the regression of the Müllerian ducts in the male embryo, and in gonadal functions in the adult. AMH is also known to be a marker of granulosa and Sertoli cell tumours. We selected a high-affinity monoclonal antibody, mAb 12G4, specific for human AMHR-II (hAMHR-II), by FACS analysis, Western blotting and immunohistochemical staining of a hAMHR-II-transfected CHO (Chinese hamster ovary) cell line, normal adult testicular tissue and granulosa cell tumours. Using peptide array screening, we identified the binding sequences of mAb 12G4 and AMH on the receptor. Identification of Asp53 and Ala55 as critical residues in the DRAQVEM minimal epitopic sequence of mAb 12G4 definitively accounted for the lack of cross-reactivity with the murine receptor, in which there is a glycine residue in place of an aspartic acid residue. In a structural model, the AMH-binding interface was mapped to the concave side of hAMHR-II, whereas the mAb 12G4-binding site was located on the convex side. mAb 12G4, the first mAb to be raised against hAMHR-II, therefore has unique properties that could make it a valuable tool for the immunotargeting of tumours expressing this receptor.

Next-Generation Genotyping by Digital PCR to Detect and Quantify the BRAF V600E Mutation in Melanoma Biopsies.

The detection of the BRAF V600E mutation in melanoma samples is used to select patients who should respond to BRAF inhibitors. Different techniques are routinely used to determine BRAF status in clinical samples. However, low tumor cellularity and tumor heterogeneity can affect the sensitivity of somatic mutation detection. Digital PCR (dPCR) is a next-generation genotyping method that clonally amplifies nucleic acids and allows the detection and quantification of rare mutations. Our aim was to evaluate the clinical routine performance of a new dPCR-based test to detect and quantify BRAF mutation load in 47 paraffin-embedded cutaneous melanoma biopsies. We compared the results obtained by dPCR with high-resolution melting curve analysis and pyrosequencing or with one of the allele-specific PCR methods available on the market. dPCR showed the lowest limit of detection. dPCR and allele-specific amplification detected the highest number of mutated samples. For the BRAF mutation load quantification both dPCR and pyrosequencing gave similar results with strong disparities in allele frequencies in the 47 tumor samples under study (from 0.7% to 79% of BRAF V600E mutations/sample). In conclusion, the four methods showed a high degree of concordance. dPCR was the more-sensitive method to reliably and easily detect mutations. Both pyrosequencing and dPCR could quantify the mutation load in heterogeneous tumor samples.

Hydrogen peroxide generation in caco-2 cell culture medium by addition of phenolic compounds: effect of ascorbic acid.

Phenolic compounds have recently attracted special attention due to their beneficial health effects; their intestinal absorption and bioavailability need, therefore, to be investigated and Caco-2 cell culture model appeared as a promising tool. We have shown herein that the addition of a grape seed extract (GSE) to Dulbecco's modified Eagle's medium (DMEM) used for Caco-2 cell culture leads to a substantial loss of catechin, epicatechin and B2 and B3 dimers from GSE in the medium after 24 h and to a production of hydrogen peroxide (H2O2). When 1420 microM ascorbic acid is added to the DMEM, such H2O2 production was prevented. This hydrogen peroxide generation substantially involves inorganic salts from the DMEM. We recommend that ascorbic acid be added to circumvent such a risk.

[Evaluation of (-2)proPSA in combination with total PSA and free PSA for the early detection of prostate cancer]. / Évaluation du test (-2)proPSA combiné au PSA total et PSA libre pour la détection précoce du cancer de la prostate.

The analytical and clinical validation of new biomarkers for the early detection of prostate is necessary. (-2)proPSA, total PSA and free PSA values are used to calculate a standardized PHI index linked to a higher probability of a positive biopsy in patients with PSA levels between 3-4 and 10 ng/L, the gray zone for prostate cancer diagnosis. The purpose of this study is to validate the analytical performance of the (-2)proPSA and to determine the predictive value of PHI for the early detection of prostate cancer. Analytical performances are correct. It is not necessary to dilute samples before analysis. The stability of (-2)proPSA is good until at least 3 hours at room temperature before centrifugation. The study of the PSAT, PSAL, (-2)proPSA and PHI values in a population of patients consulting for an early prostate cancer diagnosis shows that the index PHI is the most powerful predictive marker of cancer with an area under ROC curve of 0.70, whereas it is only 0.56 for total PSA.

[Iron metabolism in breast cancer: knowledge and future]. / Métabolisme du fer et cancer du sein: connaissances et perspectives.

Iron plays a fundamental role in biology and its concentration in living organisms is regulated very precisely. Many molecules of storage and transportation are used to maintain the intracellular homeostasis. Cancer cells have alterations in this balance. Recent studies have shown that breast cancer cells present abnormal expression of several proteins such as hepcidin and ferroportin. A prognostic impact of these alterations has been reported in patients with breast cancer. Regulatory molecules of iron metabolism could become therapeutic targets. This is an innovative approach that has emerged for treating a cancer which, despite advances in treatment and the emergence of targeted therapies, remains the leading cause of cancer death in women.