VIRGO: visualization of A-to-I RNA editing sites in genomic sequences.

RNA Editing is a type of post-transcriptional modification that takes place in the eukaryotes. It alters the sequence of primary RNA transcripts by deleting, inserting or modifying residues. Several forms of RNA editing have been discovered including A-to-I, C-to-U, U-to-C and G-to-A. In recent years, the application of global approaches to the study of A-to-I editing, including high throughput sequencing, has led to important advances. However, in spite of enormous efforts, the real biological mechanism underlying this phenomenon remains unknown.In this work, we present VIRGO (http://atlas.dmi.unict.it/virgo webcite), a web-based tool that maps Ato-G mismatches between genomic and EST sequences as candidate A-to-I editing sites. VIRGO is built on top of a knowledge-base integrating information of genes from UCSC, EST of NCBI, SNPs, DARNED, and Next Generations Sequencing data. The tool is equipped with a user-friendly interface allowing users to analyze genomic sequences in order to identify candidate A-to-I editing sites

MicroRNA fingerprints in juvenile myelomonocytic leukemia (JMML) identified miR-150-5p as a tumor suppressor and potential target for treatment

Juvenile myelomonocytic leukemia (JMML) is an aggressive leukemia of early childhood characterized by aberrant proliferation of myelomonocytic cells and hypersensitivity to GM-CSF stimulation. Mutually exclusive mutations in the RAS/ERK pathway genes such as PTPN11, NRAS, KRAS, CBL, or NF1 are found in ~90% of the cases. These mutations give rise to disease at least in part by activating STAT5 through phosphorylation and by promoting cell growth. MicroRNAs (miRs) are small non-coding RNAs that regulate gene expression, which are often deregulated in leukemia. However, little is known about their role in JMML. Here, we report distinctive miR expression signatures associated with the molecular subgroups of JMML. Among the downregulated miRs in JMML, miR-150-5p was found to target STAT5b, a gene which is often over-activated in JMML, and contributes to the characteristic aberrant signaling of this disorder. Moreover, loss of miR-150-5p and upregulation of STAT5b expression were also identified in a murine model of JMML. Ectopic overexpression of miR-150-5p in mononuclear cells from three JMML patients significantly decreased cell proliferation. Altogether, our data indicate that miR expression is deregulated in JMML and may play a role in the pathogenesis of this disorder by modulating key effectors of cytokine receptor pathways

Failure of imaging techniques in revealing breast cancer progression

This study focuses on a case of a 67-year-old woman with occult breast cancer involving the axillary lymph nodes. The instrumental examinations employed, positron emission tomography included, were not useful in diagnosing the disease. When the patient was surgically treated micro-invasive breast cancer was diagnosed. This peculiar malignant pathology is a matter of discussion especially because it is hardly diagnosable. Because of such diagnostic difficulties it may happen that micro-invasive carcinoma progression can easily mislead routine diagnostic screenings performed on women over 50

VIRES: visualization and identification of A-to-I RNA editing sites in genomic sequences

Motivations. RNA Editing is a type of post-transcriptional modification that takes place in the eukaryotes and represents one of the last frontiers of molecular biology. It alters the sequence of primary RNA transcripts by deleting, inserting or modifying residues. Several forms of RNA editing have been discovered including A-to-I, C-to-U, U-to-C and G-to-A editing. A-to-I RNA editing (Adenosine-to-Inosine) is the most frequent and common post-transcriptional modification, where adenosine (A) deamination produces its conversion into inosine (I), which in turn is interpreted by the machinery translation and splicing as guanosine (G) and so this causes the change of the RNA sequence. This biological phenomenon is catalyzed by members of the Adenosine Deaminase Acting on RNA (ADAR) family of enzymes and occurs only on dsRNA structures. Thus, A-to-I editing changes RNA molecules in various ways including: the translation of its codons; the creation and/or destruction of splicing sites; the microRNA/mRNA binding. Therefore, it is not surprising that the malfunction of the editing machinery has been implicated in various human diseases. In the last years, the application of global approaches to the study of A-to-I editing, including high throughput sequencing and bioinformatics, has led to important advances. However, in spite of enormous efforts, the real biological functioning of this phenomenon remains unknown. In this work, starting from genomic sequences, given as input, we present a bioinformatics approach to discover and visualize A-to-I editing sites. Methods. VIRES is a web-based tool that maps newly predicted and known A-to-I editing site in genomic sequences. The tool is equipped with a user-friendly interface allowing users to analyze genomic sequences in order to identify candidate A-to-I editing sites. VIRES action can be subdivided in two different tasks: the identification of known editing sites and the mapping of new ones. The system highlights the known editing events falling into the input sequences by searching the genomic positions of sequences in the DARNED dataset containing approximately 42,000 human genome loci corresponding to validated A-to-I RNA editing sites. In the second phase, we search for new putative editing events in these sequences. This task is performed by analyzing more than 38,000 Human genes (build GRCCh37/hg19). In more details, the searching of new editing sites can be divided into the following steps. First, we execute BLAST between Human genes and EST (Expressed Sequence Tag) sequences selecting only the alignments which contains at least one A-G mismatch between genomic and EST sequence. Next, we remove all mismatches that are SNPs (Single Nucleotide Polymorphisms). Finally, we verify if this candidate edited site belongs to a double strand region. Once we identify the candidate A-to-I editing sites, we add the information for the functional enrichment including the presence of repetitive elements, the ESTs sequences with the candidate editing sites, and the location of two novel motifs characterizing A-to-I editing events (CCAG[G|C]CTGG and CTG[T|G][G|A]AT[C|T][A|C]CAG) in flanking regions of putative editing sites. The user can choose whether to download this information in a text or xml file. Results. VIRES is an easy to use bioinformatic tool that allows the in-silico-identification of putative and validated A-to-I RNA editing sites. It is built on top of several knowledge bases such as DARNED, EST, SNPs

Mitomycin C and etoposide in advanced colorectal carcinoma. A clinical and in vitro experience that focuses the problem of schedule dependence in combination therapy

Abstract Background: Aim of this study was to evaluate the activity of a combination regimen of chemotherapy containing mitomycin C (MMC) and etoposide (ETO) in advanced colorectal carcinoma. Methods: Fourteen pretreated patients received MMC 2 mg/m2 and ETO 60 mg/m2, days 1–5 every 28 days. The clinical study was interrupted since no clinical response was observed in 14 patients following four courses of chemotherapy. An in vitro study was then performed on HTC-8 cell line. The cytotoxic activity of the MMC/ETO combination was tested by sulforhodamine B assay and the type of drug interaction was assessed using the method of Chou and Talalay. Cell cycle perturbations and apoptosis were evaluated by flow cytometry. Results: While MMC and ETO were singularly active, the simultaneous exposure of cells to both drugs and the sequence MMC?ETO ensued in antagonistic interaction at all levels of killed cell fraction. Conversely, the sequence ETO?MMC produced a synergistic interaction. Conclusions: These results suggest that the activity of the MMC/ETO combination is highly schedule-dependent and that the experimental drug associations should be based on a preclinical rationale before clinical trials are designed

MiR-124a regulates extracellular vesicle release by targeting GTPase rabs in lung cancer

Lung cancer is the leading cause of cancer mortality worldwide. Increased understanding of the molecular mechanisms of the disease has led to the development of novel therapies and improving outcomes. Recently, extracellular vesicles (EVs) have emerged as vehicles for the transfer of genetic information between tumors and their microenvironment and have been implicated in lung cancer initiation, progression, and response to therapy. However, the mechanisms that drive the biogenesis and selective packaging of EVs remain poorly understood. Rab family guanosine triphosphates (GTPases) and their regulators are important membrane trafficking organizers. In this study, we investigated the role of select Rab GTPases on the regulation of EV release. We found that microRNAs target Rab GTPases to regulate EV release from lung cancer cell lines. In particular, Rab32 is a target of miR-124a, and siRNA and miRNA mediated inhibition of Rab32 leads to impaired EV secretion. The downstream implications for microRNA-based regulation of EV release are currently under investigation

Non-coding RNA editing in cancer pathogenesis

In the last two decades, RNA post-transcriptional modifications, including RNA editing, have been the subject of increasing interest among the scientific community. The efforts of the Human Genome Project combined with the development of new sequencing technologies and dedicated bioinformatic approaches created to detect and profile RNA transcripts have served to further our understanding of RNA editing. Investigators have determined that non-coding RNA (ncRNA) A-to-I editing is often deregulated in cancer. This discovery has led to an increased number of published studies in the field. However, the eventual clinical application for these findings remains a work in progress. In this review, we provide an overview of the ncRNA editing phenomenon in cancer. We discuss the bioinformatic strategies for RNA editing detection as well as the potential roles for ncRNA A to I editing in tumor immunity and as clinical biomarkers