Enrica Calura

Association between miR-200c and the survival of patients with stage I epithelial ovarian cancer: a retrospective study of two independent tumour tissue collections.

BACKGROUND International Federation of Gynecology and Obstetrics stage I epithelial ovarian cancer (EOC) has a significantly better prognosis than stage III/IV EOC, with about 80% of patients surviving at 5 years (compared with about 20% of those with stage III/IV EOC). However, 20% of patients with stage I EOC relapse within 5 years. It is therefore crucial that the biological properties of stage I EOCs are further elucidated. MicroRNAs (miRNAs) have shown diagnostic and prognostic potential in stage III and IV EOCs, but the small number of patients diagnosed with stage I EOC has so far prevented an investigation of its molecular features. We profiled miRNA expression in stage I EOC tumours to assess whether there is a miRNA signature associated with overall and progression-free survival (PFS) in stage I EOC. METHODS We analysed tumour samples from 144 patients (29 of whom relapsed) with stage I EOC gathered from two independent tumour tissue collections (A and B), both with a median follow-up of 9 years. 89 samples from tumour tissue collection A were stratified into a training set (51 samples, 15 of which were from patients who relapsed) for miRNA signature generation, and into a validation set (38 samples, seven of which were from patients who relapsed) for signature validation. Tumour tissue collection B (55 samples, seven of which were from patients who relapsed) was used as an independent test set. The Cox proportional hazards model and the log-rank test were used to assess the correlation of quantitative reverse transcription PCR (qRT-PCR)-validated miRNAs with overall survival and PFS. FINDINGS A signature of 34 miRNAs associated with survival was generated by microarray analysis in the training set. In both the training set and validation set, qRT-PCR analysis confirmed that 11 miRNAs (miR-214, miR-199a-3p, miR-199a-5p, miR-145, miR-200b, miR-30a, miR-30a*, miR-30d, miR-200c, miR-20a, and miR-143) were expressed differently in relapsers compared with non-relapsers. Three of these miRNAs (miR-200c, miR-199a-3p, miR-199a-5p) were associated with PFS, overall survival, or both in multivariate analysis. qRT-PCR analysis in the test set confirmed the downregulation of miR-200c in relapsers compared with non-relapsers, but not the upregulation of miR-199a-3p and miR-199a-5p. Multivariate analysis confirmed that downregulation of miR-200c in the test set was associated with overall survival (HR 0·094, 95% CI 0·012-0·766, p=0·0272) and PFS (0·035, 0·004-0·311; p=0·0026), independent of clinical covariates. INTERPRETATION miR-200c has potential as a predictor of survival, and is a biomarker of relapse, in stage I EOC. FUNDING Nerina and Mario Mattioli Foundation, Cariplo Foundation (Grant Number 2010-0744), and the Italian Association for Cancer Research.

graphite - a Bioconductor package to convert pathway topology to gene network

BackgroundGene set analysis is moving towards considering pathway topology as a crucial feature. Pathway elements are complex entities such as protein complexes, gene family members and chemical compounds. The conversion of pathway topology to a gene/protein networks (where nodes are a simple element like a gene/protein) is a critical and challenging task that enables topology-based gene set analyses.Unfortunately, currently available R/Bioconductor packages provide pathway networks only from single databases. They do not propagate signals through chemical compounds and do not differentiate between complexes and gene families.ResultsHere we present graphite, a Bioconductor package addressing these issues. Pathway information from four different databases is interpreted following specific biologically-driven rules that allow the reconstruction of gene-gene networks taking into account protein complexes, gene families and sensibly removing chemical compounds from the final graphs. The resulting networks represent a uniform resource for pathway analyses. Indeed, graphite provides easy access to three recently proposed topological methods. The graphite package is available as part of the Bioconductor software suite.Conclusionsgraphite is an innovative package able to gather and make easily available the contents of the four major pathway databases. In the field of topological analysis graphite acts as a provider of biological information by reducing the pathway complexity considering the biological meaning of the pathway elements.

Adhesion to carbon nanotube conductive scaffolds forces action-potential appearance in immature rat spinal neurons.

In the last decade, carbon nanotube growth substrates have been used to investigate neurons and neuronal networks formation in vitro when guided by artificial nano-scaled cues. Besides, nanotube-based interfaces are being developed, such as prosthesis for monitoring brain activity. We recently described how carbon nanotube substrates alter the electrophysiological and synaptic responses of hippocampal neurons in culture. This observation highlighted the exceptional ability of this material in interfering with nerve tissue growth. Here we test the hypothesis that carbon nanotube scaffolds promote the development of immature neurons isolated from the neonatal rat spinal cord, and maintained in vitro. To address this issue we performed electrophysiological studies associated to gene expression analysis. Our results indicate that spinal neurons plated on electro-conductive carbon nanotubes show a facilitated development. Spinal neurons anticipate the expression of functional markers of maturation, such as the generation of voltage dependent currents or action potentials. These changes are accompanied by a selective modulation of gene expression, involving neuronal and non-neuronal components. Our microarray experiments suggest that carbon nanotube platforms trigger reparative activities involving microglia, in the absence of reactive gliosis. Hence, future tissue scaffolds blended with conductive nanotubes may be exploited to promote cell differentiation and reparative pathways in neural regeneration strategies.

Graphite Web: web tool for gene set analysis exploiting pathway topology

Graphite web is a novel web tool for pathway analyses and network visualization for gene expression data of both microarray and RNA-seq experiments. Several pathway analyses have been proposed either in the univariate or in the global and multivariate context to tackle the complexity and the interpretation of expression results. These methods can be further divided into ‘topological’ and ‘non-topological’ methods according to their ability to gain power from pathway topology. Biological pathways are, in fact, not only gene lists but can be represented through a network where genes and connections are, respectively, nodes and edges. To this day, the most used approaches are non-topological and univariate although they miss the relationship among genes. On the contrary, topological and multivariate approaches are more powerful, but difficult to be used by researchers without bioinformatic skills. Here we present Graphite web, the first public web server for pathway analysis on gene expression data that combines topological and multivariate pathway analyses with an efficient system of interactive network visualizations for easy results interpretation. Specifically, Graphite web implements five different gene set analyses on three model organisms and two pathway databases. Graphite Web is freely available at http://graphiteweb.bio.unipd.it/.

Analysis of Differential miRNA Expression in Primary Tumor and Stroma of Colorectal Cancer Patients

Microarray technology was used to profile miRNA expression in primary tumor and stromal tissue from paraffin embedded material of 51 patients with colorectal cancer. 26 miRNAs resulted differentially expressed with at least 2-fold change in tumor tissue with respect to stroma (16 more expressed in the tumor and 10 more expressed in the stroma). 10/26 were confirmed as differentially expressed at qRTPCR: miR-200c-3p, miR-141-3p, miR-200b-3p, miR-200a-3p, miR-1246, miR-92a-3p, miR-194-5p, miR-192-5p, miR-3651-5p, and miR-574-3p. No significant association was found between miRNA expressions and stage at diagnosis, site of primary tumor, first site of metastasis, progression-free, or overall survival.

miRNA Landscape in Stage I Epithelial Ovarian Cancer Defines the Histotype Specificities

Purpose: Epithelial ovarian cancer (EOC) is one of the most lethal gynecologic diseases, with survival rate virtually unchanged for the past 30 years. EOC comprises different histotypes with molecular and clinical heterogeneity, but up till now the present gold standard platinum-based treatment has been conducted without any patient stratification. The aim of the present study is to generate microRNA (miRNA) profiles characteristic of each stage I EOC histotype, to identify subtype-specific biomarkers to improve our understanding underlying the tumor mechanisms. Experimental Design: A collection of 257 snap-frozen stage I EOC tumor biopsies was gathered together from three tumor tissue collections and stratified into independent training (n = 183) and validation sets (n = 74). Microarray and quantitative real-time PCR (qRT-PCR) were used to generate and validate the histotype-specific markers. A novel dedicated resampling inferential strategy was developed and applied to identify the highest reproducible results. mRNA and miRNA profiles were integrated to identify novel regulatory circuits. Results: Robust miRNA markers for clear cell and mucinous histotypes were found. Specifically, the clear cell histotype is characterized by a five-fold (log scale) higher expression of miR-30a and miR-30a*, whereas mucinous histotype has five-fold (log scale) higher levels of miR-192/194. Furthermore, a mucinous-specific regulatory loop involving miR-192/194 cluster and a differential regulation of E2F3 in clear cell histotype were identified. Conclusions: Our findings showed that stage I EOC histotypes have their own characteristic miRNA expression and specific regulatory circuits. Clin Cancer Res; 19(15); 4114–23. ©2013 AACR.

The Biological Connection Markup Language

Motivation: Many models and analysis of signaling pathways have been proposed. However, neither of them takes into account that a biological pathway is not a fixed system, but instead it depends on the organism, tissue and cell type as well as on physiological, pathological and experimental conditions. Results: The Biological Connection Markup Language (BCML) is a format to describe, annotate and visualize pathways. BCML is able to store multiple information, permitting a selective view of the pathway as it exists and/or behave in specific organisms, tissues and cells. Furthermore, BCML can be automatically converted into data formats suitable for analysis and into a fully SBGN-compliant graphical representation, making it an important tool that can be used by both computational biologists and ‘wet lab’ scientists. Availability and implementation: The XML schema and the BCML software suite are freely available under the LGPL for download at http://bcml.dc-atlas.net. They are implemented in Java and supported on MS Windows, Linux and OS X. Contact: duccio.cavalieri@unifi.it; sorin@wayne.edu Supplementary information: Supplementary data are available at Bioinformatics online.

Wiring miRNAs to pathways: a topological approach to integrate miRNA and mRNA expression profiles

The production rate of gene expression data is nothing less than astounding. However, with the benefit of hindsight we can assert that, since we completely ignored the non-coding part of the transcriptome, we spent the last decade to study cell mechanisms having few data in our hands. In this scenario, microRNAs, which are key post-trascriptional regulators, deserve special attention. Given the state of knowledge about their biogenesis, mechanisms of action and the numerous experimentally validated target genes, miRNAs are also gradually appearing in the formal pathway representations such as KEGG and Reactome maps. However, the number of miRNAs annotated in pathway maps are very few and pathway analyses exploiting this new regulatory layer are still lacking. To fill these gaps, we present ‘micrographite’ a new pipeline to perform topological pathway analysis integrating gene and miRNA expression profiles. Here, micrographite is used to study and dissect the epithelial ovarian cancer gene and miRNA transcriptome defining and validating a new regulatory circuit related to ovarian cancer histotype specificity.

DC-ATLAS: a systems biology resource to dissect receptor specific signal transduction in dendritic cells

BackgroundThe advent of Systems Biology has been accompanied by the blooming of pathway databases. Currently pathways are defined generically with respect to the organ or cell type where a reaction takes place. The cell type specificity of the reactions is the foundation of immunological research, and capturing this specificity is of paramount importance when using pathway-based analyses to decipher complex immunological datasets. Here, we present DC-ATLAS, a novel and versatile resource for the interpretation of high-throughput data generated perturbing the signaling network of dendritic cells (DCs).ResultsPathways are annotated using a novel data model, the Biological Connection Markup Language (BCML), a SBGN-compliant data format developed to store the large amount of information collected. The application of DC-ATLAS to pathway-based analysis of the transcriptional program of DCs stimulated with agonists of the toll-like receptor family allows an integrated description of the flow of information from the cellular sensors to the functional outcome, capturing the temporal series of activation events by grouping sets of reactions that occur at different time points in well-defined functional modules.ConclusionsThe initiative significantly improves our understanding of DC biology and regulatory networks. Developing a systems biology approach for immune system holds the promise of translating knowledge on the immune system into more successful immunotherapy strategies.

Disentangling the microRNA regulatory milieu in multiple myeloma: integrative genomics analysis outlines mixed miRNA-TF circuits and pathway-derived networks modulated in t(4;14) patients

The identification of overexpressed miRNAs in multiple myeloma (MM) has progressively added a further level of complexity to MM biology. miRNA and gene expression profiles of two large representative MM datasets, available from retrospective and prospective series and encompassing a total of 249 patients at diagnosis, were analyzed by means of in silico integrative genomics methods, based on MAGIA2 and Micrographite computational procedures. We first identified relevant miRNA/transcription factors/target gene regulation circuits in the disease and linked them to biological processes. Members of the miR-99b/let-7e/miR-125a cluster, or of its paralog, upregulated in t(4;14), were connected with the specific transcription factors PBX1 and CEBPA and several target genes. These results were validated in two additional independent plasma cell tumor datasets. Then, we reconstructed a non-redundant miRNA-gene regulatory network in MM, linking miRNAs, such as let-7g, miR-19a, mirR-20a, mir-21, miR-29 family, miR-34 family, miR-125b, miR-155, miR-221 to pathways associated with MM subtypes, in particular the ErbB, the Hippo, and the Acute myeloid leukemia associated pathways.