Mervi Heiskanen

The MGED Ontology: a resource for semantics-based description of microarray experiments

MOTIVATION The generation of large amounts of microarray data and the need to share these data bring challenges for both data management and annotation and highlights the need for standards. MIAME specifies the minimum information needed to describe a microarray experiment and the Microarray Gene Expression Object Model (MAGE-OM) and resulting MAGE-ML provide a mechanism to standardize data representation for data exchange, however a common terminology for data annotation is needed to support these standards. RESULTS Here we describe the MGED Ontology (MO) developed by the Ontology Working Group of the Microarray Gene Expression Data (MGED) Society. The MO provides terms for annotating all aspects of a microarray experiment from the design of the experiment and array layout, through to the preparation of the biological sample and the protocols used to hybridize the RNA and analyze the data. The MO was developed to provide terms for annotating experiments in line with the MIAME guidelines, i.e. to provide the semantics to describe a microarray experiment according to the concepts specified in MIAME. The MO does not attempt to incorporate terms from existing ontologies, e.g. those that deal with anatomical parts or developmental stages terms, but provides a framework to reference terms in other ontologies and therefore facilitates the use of ontologies in microarray data annotation. AVAILABILITY The MGED Ontology version.1.2.0 is available as a file in both DAML and OWL formats at http://mged.sourceforge.net/ontologies/index.php. Release notes and annotation examples are provided. The MO is also provided via the NCICBs Enterprise Vocabulary System (http://nciterms.nci.nih.gov/NCIBrowser/Dictionary.do). CONTACT Stoeckrt@pcbi.upenn.edu SUPPLEMENTARY INFORMATION Supplementary data are available at Bioinformatics online.

Cloning of BCAS3 (17q23) and BCAS4 (20q13) genes that undergo amplification, overexpression, and fusion in breast cancer†

In breast cancer, several chromosomal sites frequently undergo amplification, implicating the location of genes important for tumor development and progression. Here we cloned two novel genes, breast carcinoma amplified sequence 3 (BCAS3) and 4 (BCAS4), from the two most common amplification sites in breast cancer, 17q23 and 20q13. The BCAS3 gene at 17q23 spans more than 600 kb at the genomic level and was predicted to encode a 913 amino acid nuclear protein. The BCAS4 gene at 20q13.2 encodes a 211 amino acid cytoplasmic protein. Both BCAS3 and BCAS4 represent novel genes with no homologies to any other known gene or protein. In the MCF7 breast cancer cell line, the BCAS3 and BCAS4 genes were co‐amplified, and cloning of a highly overexpressed 1.3‐kb transcript revealed a rearrangement fusing the last two exons of BCAS3 with BCAS4. The fusion led to a novel message in which only the first exon of BCAS4 and part of exon 23 of BCAS3 were transcribed. The BCAS4–BCAS3 fusion transcript was detected only in MCF7 cells, but the BCAS4 gene was also overexpressed in nine of 13 breast cancer cell lines. In conclusion, our results indicate that these novel genes, BCAS3 at 17q23 and BCAS4 at 20q13.2, undergo amplification, overexpression, and fusion in breast cancer and therefore may have a role in the frequent chromosomal alterations affecting these two loci. Published 2002 Wiley‐Liss, Inc.

FIBER-FISH : EXPERIENCES AND A REFINED PROTOCOL

One of the most time-consuming steps in positional cloning is the physical mapping of probes from the critical chromosomal region and the assembly of a genomic contig of large insert probes. New high-resolution Fiber-FISH techniques have significantly facilitated this tedious task by enabling rapid direct visualization of the order, degree of overlap and gap sizes of adjacent large insert clones. We have developed a method, where agarose-embedded DNA (PFGE block) is used as a source for preparing linearized DNA targets on microscope slides. This modification of the fiber-FISH technique has been successfully used in physical mapping in the 1-300 kb range as well as for detecting genomic rearrangements. Here, we present a refined protocol of our original technique. The application of this technique to agarose embedded yeast cells is also demonstrated. Finally, critical steps and trouble shooting of the method are addressed.

CGH, cDNA and Tissue Microarray Analyses Implicate FGFR2 Amplification in a Small Subset of Breast Tumors

Multiple regions of the genome are often amplified during breast cancer development and progression, as evidenced in a number of published studies by comparative genomic hybridization (CGH). However, only relatively few target genes for such amplifications have been identified. Here, we indicate how small‐scale commercially available cDNA and CGH microarray formats combined with the tissue microarray technology enable rapid identification of putative amplification target genes as well as analysis of their clinical significance. According to CGH, the SUM‐52 breast cancer cell line harbors several high‐level DNA amplification sites, including the 10q26 chromosomal region where the fibroblast growth factor receptor 2 (FGFR2) gene has been localized. High level amplification of FGFR2 in SUM‐52 was identified using CGH analysis on a microarray of BAC clones. A cDNA microarray survey of 588 genes showed >40‐fold overexpression of FGFR2. Finally, a tissue microarray based FISH analysis of 750 uncultured primary breast cancers demonstrated in vivo amplification of the FGFR2 gene in about 1% of the tumors. In conclusion, three consecutive microarray (CGH, cDNA and tissue) experiments revealed high‐level amplification and overexpression of the FGFR2 in a breast cancer cell line, but only a low frequency of involvement in primary breast tumors. Applied to a genomic scale with larger arrays, this strategy should facilitate identification of the most important target genes for cytogenetic rearrangements, such as DNA amplification sites detected by conventional CGH. Figures on http://www.esacp.org/acp/2001/22‐4/heiskanen.htm

The Ontology for Biomedical Investigations

The Ontology for Biomedical Investigations (OBI) is an ontology that provides terms with precisely defined meanings to describe all aspects of how investigations in the biological and medical domains are conducted. OBI re-uses ontologies that provide a representation of biomedical knowledge from the Open Biological and Biomedical Ontologies (OBO) project and adds the ability to describe how this knowledge was derived. We here describe the state of OBI and several applications that are using it, such as adding semantic expressivity to existing databases, building data entry forms, and enabling interoperability between knowledge resources. OBI covers all phases of the investigation process, such as planning, execution and reporting. It represents information and material entities that participate in these processes, as well as roles and functions. Prior to OBI, it was not possible to use a single internally consistent resource that could be applied to multiple types of experiments for these applications. OBI has made this possible by creating terms for entities involved in biological and medical investigations and by importing parts of other biomedical ontologies such as GO, Chemical Entities of Biological Interest (ChEBI) and Phenotype Attribute and Trait Ontology (PATO) without altering their meaning. OBI is being used in a wide range of projects covering genomics, multi-omics, immunology, and catalogs of services. OBI has also spawned other ontologies (Information Artifact Ontology) and methods for importing parts of ontologies (Minimum information to reference an external ontology term (MIREOT)). The OBI project is an open cross-disciplinary collaborative effort, encompassing multiple research communities from around the globe. To date, OBI has created 2366 classes and 40 relations along with textual and formal definitions. The OBI Consortium maintains a web resource (http://obi-ontology.org) providing details on the people, policies, and issues being addressed in association with OBI. The current release of OBI is available at http://purl.obolibrary.org/obo/obi.owl.

Identification of YAC clones for human chromosome 1p32 and physical mapping of the infantile neuronal ceroid lipofuscinosis (INCL) locus.

Infantile neuronal ceroid lipofuscinosis (INCL, CLN1) is a neurodegenerative disorder in which the biochemical defect is unknown. We earlier assigned the disease locus to chromosome 1p32 in the immediate vicinity of the highly informative HY-TM1 marker by linkage and linkage disequilibrium analysis. Here we report the construction of PFGE maps on the CLN1 region covering a total of 4 Mb of this relatively poorly mapped chromosomal region. We established the order of loci at 1p32 as tel-D1S57-L-myc-HY-TM1-rlf-COL9A2-D1S193-D1S6 2-D1S211-cen by combining data obtained from analysis of a chromosome 1 somatic cell hybrid panel, PFGE, and interphase FISH. We isolated YACs and constructed two separate YAC contigs, the loci L-myc, HY-TM1, rlf, and COL9A2 being present on a 1000-kb contig and the markers D1S193, D1S62, and D1S211 on a YAC contig spanning a maximum of 860 kb. Within the 1000-kb contig we were able to identify five CpG islands in addition to those associated with the earlier cloned genes. The YAC contigs as well as the physical map provide us with tools for the identification of the INCL gene.

High-resolution Fluorescence In Situ Hybridization: A New Approach in Genome Mapping

Mapping of the human genome has been a global effort utilizing both genetic and physical mapping techniques. One approach which has greatly facilitated the physical mapping of the human genome is fluorescence in situ hybridization (FISH). Although FISH is by now a well-established technology, new recently developed modifications have enabled an easier use and higher resolution. The high-resolution FISH techniques have given a special impact in positional cloning: searching the functional gene from a chromosomal area where the gene has been genetically localized. New high-resolution FISH techniques include hybridization of probes to free chromatin, DNA fibres or mechanically stretched chromosomes. These targets have widened the resolution of FISH to detect distances from the traditional cytogenetic resolution level down to a resolution of a few kilobases. They also have significantly speeded up high-resolution physical mapping and thus made the search of new disease genes easier.

Idiopathic macrocytic anaemia in the aged: molecular and cytogenetic findings

Summary. Macrocytosis in the elderly is often caused by abnormalities of haematological stem cell differentiation. In this study, a group of elderly patients was analysed for four molecular and cell biological parameters. The aim of the study was to screen elderly patients with idiopathic macrocytic anaemia or MDS for a set of alterations which are related to haematological dysplasia. The analyses used were: DNA‐methylation at the calcitonin A gene 5′‐area, NBAS point mutations at codons 12 and 13, in vitro colony formation of peripheral blood progenitor cells and cytogenetics of bone marrow cells. The results show that a significant portion of elderly patients with idiopathic macrocytosis have one or more of the abnormalities analysed. Hypermethylation of the calcitonin A gene 5′‐area at the chromosome 11 band pi 5 is relatively common (7/15). Chromosomal aberrations (3/12) and NRAS oncogene point mutations (0/15) were rare findings. In vitro culture of erythroid progenitor cells was relatively frequently abnormal (7/15). Eight of our nine macrocytic patients who did not fulfill the FAB criteria for MDS had at least one of the alterations studied; this suggests that these patients might represent early phases of a stem cell disorder.

caNanoLab: data sharing to expedite the use of nanotechnology in biomedicine.

The use of nanotechnology in biomedicine involves the engineering of nanomaterials to act as therapeutic carriers, targeting agents and diagnostic imaging devices. The application of nanotechnology in cancer aims to transform early detection, targeted therapeutics and cancer prevention and control. To assist in expediting and validating the use of nanomaterials in biomedicine, the National Cancer Institute (NCI) Center for Biomedical Informatics and Information Technology, in collaboration with the NCI Alliance for Nanotechnology in Cancer (Alliance), has developed a data sharing portal called caNanoLab. caNanoLab provides access to experimental and literature curated data from the NCI Nanotechnology Characterization Laboratory, the Alliance and the greater cancer nanotechnology community.

Experiences in supporting the structured collection of cancer nanotechnology data using caNanoLab.

Summary The cancer Nanotechnology Laboratory (caNanoLab) data portal is an online nanomaterial database that allows users to submit and retrieve information on well-characterized nanomaterials, including composition, in vitro and in vivo experimental characterizations, experimental protocols, and related publications. Initiated in 2006, caNanoLab serves as an established resource with an infrastructure supporting the structured collection of nanotechnology data to address the needs of the cancer biomedical and nanotechnology communities. The portal contains over 1,000 curated nanomaterial data records that are publicly accessible for review, comparison, and re-use, with the ultimate goal of accelerating the translation of nanotechnology-based cancer therapeutics, diagnostics, and imaging agents to the clinic. In this paper, we will discuss challenges associated with developing a nanomaterial database and recognized needs for nanotechnology data curation and sharing in the biomedical research community. We will also describe the latest version of caNanoLab, caNanoLab 2.0, which includes enhancements and new features to improve usability such as personalized views of data and enhanced search and navigation.