Proteomic analysis of nipple aspirate fluid to detect biologic markers of breast cancer.

The early detection of breast cancer is the best means to minimise disease-related mortality. Current screening techniques have limited sensitivity and specificity. Breast nipple aspirate fluid can be obtained noninvasively and contains proteins secreted from ductal and lobular epithelia. Nipple aspirate fluid proteins are breast specific and generally more concentrated than corresponding blood levels. Proteomic analysis of 1 microl of diluted nipple aspirate fluid over a 5-40 kDa range from 20 subjects with breast cancer and 13 with nondiseased breasts identified five differentially expressed proteins. The most sensitive and specific proteins were 6500 and 15 940 Da, found in 75-84% of samples from women with cancer but in only 0-9% of samples from normal women. These findings suggest that (1) differential expression of nipple aspirate fluid proteins exists between women with normal and diseased breasts, and (2) analysis of these proteins may predict the presence of breast cancer

Transcription profiles of non-immortalized breast cancer cell lines

BACKGROUND: Searches for differentially expressed genes in tumours have made extensive use of array technology. Most samples have been obtained from tumour biopsies or from established tumour-derived cell lines. Here we compare cultures of non-immortalized breast cancer cells, normal non-immortalized breast cells and immortalized normal and breast cancer cells to identify which elements of a defined set of well-known cancer-related genes are differentially expressed. METHODS: Cultures of cells from pleural effusions or ascitic fluids from breast cancer patients (MSSMs) were used in addition to commercially-available normal breast epithelial cells (HMECs), established breast cancer cell lines (T-est) and established normal breast cells (N-est). The Atlas Human Cancer 1.2 cDNA expression array was employed. The data obtained were analysed using widely-available statistical and clustering software and further validated through real-time PCR. RESULTS: According to Significance Analysis of Microarray (SAM) and AtlasImage software, 48 genes differed at least 2-fold in adjusted intensities between HMECs and MSSMs (p < 0.01). Some of these genes have already been directly linked with breast cancer, metastasis and malignant progression, whilst others encode receptors linked to signal transduction pathways or are otherwise related to cell proliferation. Fifty genes showed at least a 2.5-fold difference between MSSMs and T-est cells according to AtlasImage, 2-fold according to SAM. Most of these classified as genes related to metabolism and cell communication. CONCLUSION: The expression profiles of 1176 genes were determined in finite life-span cultures of metastatic breast cancer cells and of normal breast cells. Significant differences were detected between the finite life-span breast cancer cell cultures and the established breast cancer cell lines. These data suggest caution in extrapolating information from established lines for application to clinical cancer research

Proteomics of breast cancer outcomes and prospects

Breast cancer is a major public health problem. The identification of new markers to differentiate neoplastic from the normal cells, more thorough understanding of different stages of the pathology, as well as the definition of new therapeutic targets, are all of critical importance. With the completion of human genome sequencing and the introduction of mass spectrometry, combined with protein identification via advanced bioinformatics, proteomics has emerged as a valuable tool for the discovery of new molecular markers. New methods in functional proteomics have also been developed to study the intracellular signaling pathways that underline the development of breast cancer. As illustrated with the examples of fibroblast growth factor-2 and H19, an oncogenic, noncoding mRNA, proteomics have become a powerful approach for deciphering the complex signaling circuitry involved in tumor growth. Breast cancer proteomics have already identified proteins of potential clinical interest (such as the molecular chaperone 14-3-3 sigma) and technological innovations in large scale/high throughput analysis are now ushering in new prospects

Enhancement of PDGF-BB mitogenic activity on human dermal fibroblasts by biospecific dextran derivatives

Dextran derivatives are biosynthetic polyanionic polymers which exert some of the heparin properties such as regulating the activity of several heparin-binding growth factors. Based on a reproducible synthetic procedure, we have synthesized a new generation of dextran derivatives named dextran methylcarboxylate benzylamide sulfate (DMCBSu). Here we investigated the ability of a library of well-characterized DMCBSu to interact with platelet-derived growth factor-BB (PDGF-BB), which has essential roles during wound healing. Using gel mobility shift assay, our results indicate that benzylamide and sulfate groups act synergically to bind to PDGF-BB. Furthermore we show that depending on their chemical composition, functionalized dextrans are able to potentiate the mitogenic activity of PDGF-BB on human dermal fibroblasts. This enhancing effect is accompanied with changes in PDGF-BB-induced signaling events, as determined by the use of specific inhibitors and by western blot. Our results suggest that the use of such biopolymers combined with a local administration of the growth factor could increase the efficiency of the biomolecule activity in future therapeutic strategies

Proteomic analysis reveals that 14-3-3 sigma in downregulated in human breast cancer cells

The class of molecular chaperones known as 14-3-3 is involved in the control of cellular growth by virtue of its apparent regulation of various signaling pathways, including the Raf/mitogen-activated protein kinase pathway. In breast cancer cells, the sigma form of 14-3-3 has been shown to interact with cyclin-dependent kinases and to control the rate of entry into mitosis. To test for a direct role for 14-3-3 in breast epithelial cell neoplasia, me have quantitated 14-3-3 protein levels using a proteomic approach based on two-dimensional electrophoresis and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF). We show here that 14-3-3 sigma protein is strongly down-regulated in the prototypic breast cancer cell lines MCF-7 and MDA-MB-231 and in primary breast carcinomas as compared with normal breast epithelial cells. In contrast, levels of the alpha, beta, delta, or zeta isoforms of 14-3-3 mere the same in both normal and transformed cells. The data support the idea that 14-3-3 sigma is involved in the neoplastic transition of breast epithelial cells by virtue of its role as a tumor suppressor; as such, it may constitute a robust marker with clinical efficacy for this pathology

Proteomic characterization in the hippocampus of prenatally stressed rats

Rats exposed to early life stress are considered as a valuable model for the study of epigenetic programming leading to mood disorders and anxiety in the adult life. Rats submitted to prenatal restraint stress (PRS) are characterized by an anxious/depressive phenotype associated with neuroadaptive changes in the hippocampus. We used the model of PRS to identify proteins that are specifically affected by early life stress. We therefore performed a proteomic analysis in the hippocampus of adult male PRS rats. We found that PRS induced changes in the expression profile of a number of proteins, involved in the regulation of signal transduction, synaptic vesicles, protein synthesis, cytoskeleton dynamics, and energetic metabolism. Immunoblot analysis showed significant changes in the expression of proteins, such as LASP-1, fascin, and prohibitin, which may lie at the core of the developmental programming triggered by early life stress. (C) 2011 Elsevier B.V. All rights reserved