Upcoming MicroRNA Focused Conference

Don’t miss the upcoming – Third International MicroRNAs Europe 2008 Meeting – MicroRNAs: Biology to Development and Disease

November 3 – 4, 2008
Venue: Peterhouse, University of Cambridge, Cambridge, UK

A Unique Theme Conference & Exhibition in the MicroRNomics!

The Venue Peterhouse (middle) is the oldest College of the University of Cambridge, founded in 1284 and an institution dedicated to education and research. Throughout its history, Petreans have been at the heart of the political, social and religious controversies that have shaped today’s society. Some of the influential Petreans include: Henry Cavendish, Lord Kelvin, Sir Frank Whittle and Christopher Cockerill, and Nobel Laureates – Sir John Kendrew, Sir Aaron Klug, Archer Martin and Max Perutz , who gave a twentieth century lead in the field of Molecular Biology. Since many centuries it remained as a hub for innovation and successive generations of the brightest young people around the world.

AGENDA/SPEAKERSMonday, November 3, 2008

7:00 – 8:30 A.M: Registration Open
7:30 – 8:45 A.M: Continental breakfast
8:00 – 9:30 A.M: Technology Session I (consists of 3 lectures)

Tuesday, November 4, 20087:00 – 8:30 A.M: Registration Open
7:30 – 8:45 A.M: Continental breakfast
 

 

 

 

 

 

 

8:00 – 9:30 A.M: Technology Session II (consists of 3 lectures)
Scientific Sessions Start at 9:30 A.M and Ends at 6.00 P.M on all two days.The actual agenda will be updated. Please visit again.

Organizers: 

 

 

 

 

 

 

Krishnarao Appasani, PhD., MBA (Chair)
Founder & CEO
GeneExpression Systems, Inc. Waltham, MA USA

Sakari Kauppinen, Ph.D.
Associate Director
Santaris Pharma, Copenhagen, Denmark
Anton J. Enright, PhD.
Group Leader at European Bioinformatics Institute
Wellcome Trust Sanger Institute, Hinxton, Cambridge, United Kingdom

Eric A. Miska, PhD.
Assistant Professor at the Wellcome Trust/Cancer Research UK/Gurdon Institute
University of Cambridge, Cambridge, United Kingdom

Inaugural Speaker & Lifetime Achievement Awardee:
Prof. Sir John Walker, D.Phil., FRS
Chemistry Nobel Laureate1997
MRC Dunn Human Nutrition Unit , Cambridge, CB2 0XY, United Kingdom
Title: TBA

Keynote Speaker & MicroRNAs Innovator Awardee of 2008
Prof. Carlo M. Croce, MD.
Chair and Professor
College of Medicine and
Public Health
Ohio State University, Columbus,OH. USA
Title: MicroRNAs in Cancer

Industry Keynote speaker
Sakari Kauppinen, PhD.
Director
Santaris Pharma, Horshol, Denmark
Title: TBA

David C. Baulcombe, PhD., FRS.
Professor of Botany & Royal Society Research Professor
University of Cambridge, Cambridge, UK
Title: Short silencing RNA in plants

Mark A. Lindsay, PhD.
Reader
Machester University, Manchester, UK
Title: MiRNAs in inflammation and lung disease

Dónal O’Carroll Ph.D.
Group Leader
EMBL Monterotondo Mouse Biology Unit
Monterotondo Scalo, Italy
Title: TBA

Markus Stoffel, MD, PhD.
Professor
Swiss Federal Institute of Technology ETH
Institute for Molecular Systems Biology, Zürich, Switzerland
Title: TBA

Dr F. Gregory Wulczyn
Professor
Institute for Cell and Neurobiology
Centrum for Anatomy
Charité –
Universitätsmedizin
Berlin, Germany
Title: TBA

Iain Russell, PhD.
Applied Biosystems, Inc.,
Foster City, CA, USA
Title: TBA

Simon Mauch, PhD.
Global Marketing Team Manager
Miltenyi Biotec GmbH, Bergisch Gladbach, Germany
Title: TBA

Cristoph Eicken, PhD.
Head of Technical Services, Microarrays
LC Sciences, LLC., Houston, TX, USA
Title: Microfluidic Technology Enabling Advances in microRNA Research

www.lcsciences.com

 

Ilona Grunwald Kadow, PhD.
Principal Investigator
Max-Planck Institute of Neurobiology, Martinsried, Germany
Title: TBA

Edoardo Missiaglia, PhD.
Postdoctoral Fellow
The Institute of Cancer Research
Sutton Surrey, UK
Title: MicroRNAs in childhood rhabdomyosarcoma

Jacob Ulrik Fog, PhD.
Senior Scientist
Exiqon A/S
Vedbaek, Denmark
Title: The development of microRNA based diagnostics for cancer treatment selection

Carlos Caldas MD FACP FRCP FMedSci
Professor of Cancer Medicine
Cambridge Research Institute
Department of Oncology, University of Cambridge
Cambridge, UK
Title: TBA

Anders Lund, PhD.
Associate Professor
Biotech Research and Innovation Centre
Copenhagen, Denmark
Title: TBA

Amy Buck, PhD.
Marie Curie Research Fellow
Center for Infectious
Diseases
University of Edinburgh
Edinburgh, United Kingdom
Title: TBA

Grace Tan, PhD.
Postdoctoral Fellow in
Rheumatology
University of Pennsylvania
Philadelphia, PA, USA
Title: Expanded Functions of Recombinant Mammalian Argonaute 2, a Key Effector Protein of RNAi

Paul Kellam, PhD.
Reader
University College of London, UK
Title: Host cell and viral miRNA expression in B cell lymphomas
 

Johan Gibcus, PhD.
Postdoctoral Fellow in
Pathology
University Medical Center Groningen
Groningen, The Netherlands
Title: miRNA profiling in HL

Cesare Peschle, M.D.
Chairman of Dept. of Hematology
Istituto Superiore di Sanità
Roma, Italy
Title: MicroRNAs in cancer and hematopoiesis

Martin Fabani, Ph.D.
Postdoctoral Fellow
MRC Laboratory Of Molecular
Biology
Hills Road, Cambridge, CB2 0QH, UK

Kim Holmstrøm, Ph.D.
Group Leader, Biomarker Projects
Bioneer A/S, Hørsholm, Denmark
Title: Establishing a technology platform for next generation in situ based microRNA diagnostics

Sarah Jugurnauth
PhD Student
GlaxoSmithKline Pharmaceuticals, UK

Joost Kluiver, PhD.
Postdoctoral Fellow in
Pathology & Lab. Medicine
University Medical Center Groningen
Groningen, The Netherlands
Title: Identification of miRNAs involved in B-cell receptor induced apoptosis

William S. Marshall, PhD.
President and Chief Executive Officer
miRagen Therapeutics
Boulder, CO , USA
Title: MicroRNA manipulation as a therapeutic strategy for cardiovascular diseas
 

Elena Vigorito, PhD.
Group Leader
Babraham Institute
Lab. Of Lymphocyte Signaling & Development
Cambridge, UK
Title: miRNA regulation of the immune system

Graham Speight, PhD.
Head of Molecular Biology
Oxford Gene Technology
Sandy Lane Yarnton
Oxford OX5 1PF, UK
Title: New High-Throughput Facility Expands miRNA Profiling Opportunity

Anton J. Enright, PhD,
Group Leader
European Bioinformatics Institute
Wellcome Trust Genome Campus, Hinxton, Cambridge, UK
Title: Detection of microRNA binding and siRNA off-targets from expression data

Mrs. Sonja Vorwerk, PhD.
Febit biotech, GmbH
Heidelberg, Germany
Title: Beyond miRNAs – ncRNAs On Flexible Arrays

Ana Soares
PHD student in
Molecular Biology
University of Aveiro, Aveiro, Portugal
Title: Identification of novel zebrafish microRNAs using massive parallel DNA sequencing

Panel Members:
Justine Muir
European Patent Attorney
Chartered Patent Attorney
BioScience Patents Ltd., UK

Stefan Scholtes, PhD.
Professor of
Management Science
University of Cambridge
Cambridge, UK

 

Key Sessions:
The most-up-to-date developments will be addressed:
MicroRNAs Discovery and Biogenesis
MicroRNAs in Development
Bioinformatics of miRNAs
MicroRNAs in Virology & Diagnostics
MicroRNAs in Stem Cell Biology
MicroRNAs in Disease Biology

ALL ABSTRACTS

miRNAs in lung inflammation and disease
Mark A. Lindsay, PhD., Reader in Biopharmaceutics, National Heart and Lung Institute, Imperial College, London, United Kingdom

Inflammation is thought to be responsible for the development of lung disease including asthma, chronic obstructive pulmonary disease and pulmonary fibrosis. In this presentation, I shall present data that indicates that changes in miRNA expression are associated with lung disease and specifically, that miR-146a and miR-223 are involved in regulating acute lung inflammation. I will also discuss the potential of modulating lung miRNAs as a novel therapeutic approach.

MicroRNAs in childhood rhabdomyosarcoma
Edoardo Missiaglia, PhD. Postdoctoral Fellow, The Institute of Cancer Research, Sutton, Uinted Kingdom

Edoardo Missiaglia1, Chris Shepherd1, Prakash K. Rao2, Janet Shipley1
1 The Institute of Cancer Research, Sutton, UK; 2 Whitehead Institute for Biomedical Research, Cambridge, USA

Introduction: Rhabdomyosarcomas are soft tissue sarcomas resembling myogenic development and are one of the leading causes of paediatric cancer death.
MicroRNAs (miRNA) are recently discovered short RNA molecules that play a key role in the regulation of gene expression. Expression of miRNAs in various tumour types correlates with patient outcome and differentiation status and have demonstrable tumour suppressor and oncogenic roles. Interestingly, some miRNAs have been involved in myogenesis.

Methods: Thirty six primary rhabdomyosarcoma (RMS) tumours, 18 RMS cell lines, 7 normal muscle samples and one myoblast were screened for miRNA expression using a microarray platform containing 359 human probes. Quantitative RT-PCR based assay was employed to validate our findings. The functional role of selected miRNA differentially expressed between RMS and normal muscle was then evaluated by artificially modulating their expression in RMS cell lines.

Results: The analysis has identified 40 and 31 miRNAs over and under expressed compared to normal muscle, respectively; five of them have been confirmed by other means in a larger set of samples. A number of the identified miRNAs have been associated with muscle differentiation and tumourigenesis, whilst others are novel. Functional studies have confirmed the involvement of selected miRNAs in RMS tumourigenesis. In addition, the expression of some miRNAs was associated with overall patient survival.

Conclusions: The analysis identified several miRNAs potentially involved in rhabdomyosarcoma development as well as associated with poorer outcome. Further studies will elucidate their biological role and their potential as therapeutic targets in rhabdomyosarcomas.

Expanded Functions of Recombinant Mammalian Argonaute 2, a Key Effector Protein of RNAi
Grace S. Tan, PhD., Postdoctoral Fellow, Dept of Rheumatology, University of Pennsylvania School of Medicine, Philadelphia PA 19104, USA

Grace S. Tan, Barry Garchow and Marianthi Kiriakidou
Dept of Rheumatology, University of Pennsylvania School of Medicine, Philadelphia PA 19104, USA

microRNAs (miRNAs) are small (~22nt) noncoding RNAs that are regulating gene expression in arthropods, nematodes, mammals, plants and viruses. miRNAs are implicated in all aspects of cellular processes including embryonic development and stem cell maintenance. Perturbations in miRNA expression have also been detected in many human diseases including heart failure and cancer. miRNAs are loaded into microribonucleoprotein effector complexes (miRNPs or RISCs) and are bound to the 3’ untranslated region (3’-UTR) of target mRNAs, resulting in translational repression or degradation of their targets. Argonaute is the key protein component of the miRNPs. In humans, there are four isoforms of Argonaute (Ago1-4), all of which are involved in miRNA mediated translational repression. Ago2 is the only human Ago protein with the additional ability to cleave target mRNAs through an RNAse H-like domain. Ago2 null mice exhibit severe cardiac and neuronal developmental defects and are embryonically lethal. We have expressed recombinant active GST-Ago2 fusion protein using the baculovirus (SF9) system. Our aim is to characterize the RNA binding and cleavage activities of the recombinant Ago2. Our results have shown that recombinant Ago2 binds strongly to small RNAs and is a multi-turnover cleavage enzyme. Interestingly, we have also shown that in vitro, recombinant Ago2 can bind and use premature miRNAs (pre-miRNAs) as guides to catalyze cleavage of perfectly complementary target RNAs. This novel interaction with pre-miRNA may give insights to undiscovered functions of Ago2.

microRNA Profiling Using a High Performance, Flexible µParaflo® Biochip Platform
Christoph Eicken, PhD., Head of Technical Services, Microarrays, LC Sciences, LLC., Houston, Texas 77054, USA

An advanced microfluidic biochip system designed to produce high quality data, stay current with the rapidly evolving microRNA field, and perform diverse small RNA discovery experiments is presented. This technology’s unique flexibility allows for miRBase synchronicity and design of customized biochips adapted to each researcher’s specific needs. Applications featuring disease marker discovery, drug treatment, microRNA target screening, and small RNA discovery are highlighted.

The development of microRNA based diagnostics for cancer treatment selection
Jacob Ulrik Fog, PhD., Senior Scientist, Exiqon A/S, Vedbaek, Denmark

Abstract:
MicroRNAs (miRNAs) are a recently discovered class of small, non-coding RNAs involved in silencing of the expression of specific genes. These miRNAs have been implicated to be important in cancer initiation and progression and have the potential to be robust biomarkers.
Using Locked Nucleic Acid (LNA) based miRNA detection technology we have developed high throughput in situ detection of miRNAs in clinical paraffin-embedded samples. These single and low complexity miRNA ISH methods give a unique insight into the localization and quantitation of miRNAs in tumors. We are developing these and LNA based QPCR as molecular diagnostic tests for treatment selection in cancer.

Host cell and viral miRNA expression in B cell lymphomas
Paul Kellam, PhD., Reader, Centre for Virology, Div. of Infection and Immunity, University College of London, London, UK

Micro RNAs (miRNAs) are a class of approximately 21- nucleotide non-coding small RNAs that post-transcriptionally down-regulate the expression of mRNAs bearing complementary target sequences. Several hundred miRNAs have been predicted within the human genome and their key role as regulators of important biological pathways such as development, proliferation, apoptosis and viral infection is becoming clear. Recent discovery of virus-encoded miRNAs, especially miRNAs encoded by herpesviruses, indicates that viruses also use this fundamental mode of gene regulation. Using Agilent miRNA microarray technology, we have developed a data new analysis method. We have determined the pattern of expression of human, Epstein Barr virus (EBV) and Kaposi’s sarcoma virus (KSHV) encoded miRNAs in B cell tumour cell lines associated with different stages of normal B cell development. Only a subset of human miRNAs are expressed in different B cell tumours cell lines and different viral miRNA expression patterns are evident in herpesvirus positive tumour types. Additionally, tumour specific down regulation of miR155, 221 and 222 occurs in KSHV infected lymphomas. These data suggest that like host and virus gene expression, host and virus miRNA expression is associated with the underlying tumour cell type.

Establishing a technology platform for next generation in situ based microRNA diagnostics
Kim Holmstrøm, PhD., Group Leader of Biomarker Projects Bioneer A/S, Hørsholm, Denmark

The in situ localization of microRNAs in diseased tissue will become an important tool for next generation diagnostic systems e.g. to provide more precise and reliable cancer diagnostics. MicroRNAs have already been established as novel and additionally informative diagnostic biomarkers in cancer. To meet the demands for coming diagnostic tools, we are currently developing a novel technology platform that will allow for multiple assays to be performed on a single tissue section by improving reaction kinetics and enabling sequential detection. Examples will be provided that illustrate the importance of in situ detection to spatially distinguish different microRNA signals, even to the level of sub-cellular localization.

 

Modulation of murine cytomegalovirus infection by host microRNAs
Amy Buck, PhD., Marie Curie Research Fellow, Center for Infectious Diseases, Division of Pathway Medicine, University of Edinburgh, Edinburgh, United KingdomMicroRNAs encoded by either host or virus can play a role viral infection, both in the acute and latent stages. We have established an in vitro system for examining the function of viral and host microRNAs on murine cytomegalovirus replication. Using this assay we have identified a number of microRNAs which modulate replication and this work is being extended to other viruses. Potential mechanisms will be discussed.
 

Advances in MicroRNA Profiling Using Quantitative Real-Time PCR
Iain Russell, PhD., Applied Biosystems, Inc., Foster City, CA, USA

Applied Biosystems Megaplex™ Primer Pools and TaqMan® MicroRNA Arrays provide an ideal workflow for the profiling of microRNAs from human, mouse and rat samples, even when only minute amounts of sample (down to 1ng) are available. Utilizing the same novel stem-loop RT primers and assay design as the individual TaqMan® MicroRNA Assays, the Megaplex™ profiling workflow is highly sensitive, offers a broad dynamic range (up to 7 logs) and is highly specific, enabling quantitation of only the mature microRNA sequences in as little as 5 hours. Here we present data demonstrating the high performance of the Megaplex™ workflow and provide specific examples of how it has been applied to real-life biological applications.

Identification of miRNAs involved in B-cell receptor induced apoptosis
Joost Kluiver, PhD., Postdoctoral Fellow in the lab of Dr. Anke van den Berg
Dept. of Pathology & Lab. Medicine, University Medical Center Groningen, Groningen, The Netherlands

Joost Kluiver1,2, Bart-Jan Kroesen1, Anke van den Berg1 and Chang-Zheng Chen2
1 Department of Pathology, University Medical Centre Groningen, Groningen, The Netherlands
2 Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, USA

B-cell receptor (BCR) induced apoptosis is of crucial importance to maintain self-tolerance. We used the WEHI-231 B-cell line as a model to study the involvement of microRNAs in BCR induced apoptosis. Expression profiling demonstrated that 33 microRNAs were differentially expressed upon BCR stimulation. Interestingly, rescue from apoptosis via CD40 costimulation almost completely reversed the expression pattern back to that of unstimulated cells. Overexpression of several candidate miRNAs in WEHI-231 cells resulted in a growth disadvantage for six miRNAs and a growth advantage for one miRNA. For two of the miRNAs whose increased expression was disadvantageous, an increased sensitivity to apoptosis was demonstrated.

miRNA profiling in Hodgkin lymphoma
Johan Gibcus, PhD., Postdoctoral Fellow in Pathology & Lab. Medicine, University Medical Center Groningen, Groningen, The Netherlands

Johan H. Gibcus1, Lu Ping Tan1, Geert Harms1, Rikst Nynke Schakel1, Debora de Jong1, Tjasso Blokzijl1, Peter Moller3, Sibrand Poppema1, Bart-Jan Kroesen2 and Anke van den Berg1

Section of Pathology1 and Section of Medical Biology2, Department Pathology & Laboratory Medicine, University Medical Center Groningen and University of Groningen, Groningen, The Netherlands. 3Institute of Pathology, University of Ulm, Ulm, Germany

Hodgkin lymphoma (HL) is derived from pre-apoptotic germinal center B-cells, although a general loss of B cell phenotype is noted. Using quantitative RT-PCR and miRNA microarray, we determined the microRNA (miRNA) profile of HL and compared this with the profile of a cell line panel of B-cell non-Hodgkin lymphomas (NHL). HL specific miRNAs included miR-17-92 cluster members, miR-16, miR-21, miR-24 and miR-155, whereas miR-150 was downregulated. We found differential expression between HL and other B-cell lymphoma derived cell lines for 27 miRNAs. Using luciferase reporter assays, we demonstrated that AGTR1, FGF7, ZNF537, ZIC3 and IKBKE are true miR-155 target genes in HL.

MicroRNA inhibition with LNA/2’-O-Methyl mixmers and Peptide Nucleic Acids
Martin Fabani, PhD. Postdoctoral Fellow in the Lab of Michael Gait, Medical Research Council Laboratory Of Molecular Biology, Hills Road, Cambridge, CB2 0QH, UK

MicroRNA knockdown by antisense oligonucleotides is a useful strategy to explore microRNA functionality and as potential therapeutics. I will present data on the application of Locked Nucleic Acids (LNA)/2’-O-Methyl mixmers and Peptide Nucleic Acid (PNA) as effective inhibitors of microRNA activity in cells, and show that microRNA inhibition can be achieved without the need for transfection by conjugation of an antisense PNA to the cell-penetrating peptide R6-Penetratin, or merely by linkage to just four Lysine residues.

let7 and Lin-28: a feedback loop controlling neural stem cell maturation
F. Gregory Wulczyn, PhD. Project Leader, Institute for Cell and Neurobiology
Centrum for Anatomy, Charité – Universitätsmedizin Berlin, Berlin, Germany

Agnieszka Rybak, Heiko Fuchs, Lena Smirnova and F. Gregory Wulczyn

miRNA populations undergo a dramatic transition during early stages of stem cell differentiation. Mammalian homologs of the paradigm miRNAs lin-4 (mir-125) and let-7, originally recognized based on mutational phenotypes in stem cell maturation, are strongly induced during neural differentiation of embryonic stem cells and embryocarcinoma cells. We have shown that neural stem cells isolated from E12 mouse cortex express let-7 as well as mir-125. Recent work has pointed to an important role for post-transcriptional mechanisms in the control of let-7 maturation during stem cell differentiation, early development and oncogenesis. We demonstrate that the pluripotency factor Lin-28 binds the pre-let-7 RNA and inhibits processing by the Dicer ribonuclease in pluripotent cells. In committed neural stem cells, Lin-28 is suppressed by mir-125 and let-7, allowing processing of pre-let-7 to proceed. Introduction of let-7 or mir-125 antagonists into neural strem cells led to upregulation of Lin-28 and loss of pre-let-7 processing activity, suggesting that let-7, mir-125 and lin-28 participate in an autoregulatory circuit to control miRNA processing during stem cell commitment.

Small RNAs, Great Challenge – Considerations for the development of microRNA microarrays
Simon Mauch, PhD., Global Marketing Team Manager MACSmolecular, Miltenyi Biotech GmbH, Bergisch Gladbach, Germany

Development of microRNA-specific microarrays is a technical challenge: the small size of microRNAs and the high sequence similarities between some microRNAs require new technical solutions to allow for significant sensitivity and specificity.
We are going to present data on the miRXplore™ microarray platform that was developed in close cooperation with leading experts at Rockefeller University(1). Reliable microRNA-specific expression profiling can be achieved using a combination of novel hybridization buffers and a proprietary hybridization technology called the active circulation.

(1) Landgraf, P. et al., Tuschl, T. (2007) Cell 129: 1401–1414.

Beyond miRNAs – ncRNAs On Flexible Arrays
Sonja Vorwerk, PhD., Febit biotech, GmbH, Heidelberg, Germany

Approximately every 3-4 months a new release of the miRBase becomes available, so analysis tools need to be flexible to incorporate these changes rapidly. In addition, there is more than just miRNAs in the field of non-coding RNAs, which are not covered by high-throughput technology at the moment. Here we present the Geniom One technology which provides the outmost flexibility for any type of ncRNA and introduce you to some of the possible applications.

Identification of novel zebrafish microRNAs using massive parallel DNA sequencing
Ana Soares, PhD student in Molecular Biology, Department of Biology, University of Aveiro, Aveiro, Portugal

Ana R. Soares1,2, Patrícia M. Pereira1, Bruno Santos3, Conceição Egas3 and Manuel A. S. Santos1,3
1Department of Biology & CESAM, University of Aveiro, 3810-193 Aveiro
2Doctoral Program of Experimental Biology and Biomedicine, Center for Neuroscience and Cell Biology, University of Coimbra, 3000 Coimbra
3Biocant Research Centre, Genomics Unit, 3060-197 Cantanhede. Portugal.

The discovery of the complete set of miRNAs in an organism is essential to understand the underlying mechanisms regulated by these small molecules. Recently massive parallel DNA sequencing has been applied successfully to miRNA finding, allowing the discovery of a significant number of novel small molecules. In this study, 454 DNA technology was used to identify new ZF miRNAs. A series of cDNA libraries were prepared from miRNAs that were isolated at different embryonic time points and from fully developed organs, to ensure a high coverage of the miRNA population in this organism. This approach retrieved 90% of the 192 known ZF miRNAs and permitted the identification of 25 novel miRNAs and many unknown miRNA star sequences. The results demonstrate that one single pyrosequencing run can be successfully used to characterize complex miRNA populations, determine their expression pattern during cell development and in adult tissues and discover novel miRNA molecules.

P53-independent up-regulation of miR-34a during oncogene-induced senescence controls central cell cycle regulators.
Anders Lund, PhD., Associate Professor, Biotech Research and
Innovation Centre
University of Copenhagen, Copenhagen, Denmark

Oncogene-induced senescence (OIS) is an important defense against tumor formation. To identify microRNAs involved in OIS we profiled microRNAs in primary human fibroblasts holding an inducible B-RAF transgene. We show that miR-34a induction during OIS is independent of p53 and identify the responsible activating transcription factor. Furthermore, we experimentally determine key downstream targets of miR-34a during OIS. The existence of multiple oncogenic pathways activating miR-34a points to important tumor-suppressor function for this microRNA.

Antagonizing microRNAs for therapeutics
Sakari Kauppinen, Ph.D., Director, Santaris Pharma, Horshol, Denmark

Sakari Kauppinen1,2

1Wilhelm Johannsen Centre of Functional Genome Research, Department of Cellular and Molecular Medicine, University of Copenhagen, Blegdamsvej 3, DK-2200, Cph N, Denmark
2Santaris Pharma, Bøge Allé 3, DK-2970 Hørsholm, Denmark

MicroRNAs (miRNAs) are an abundant class of short endogenous non-coding RNAs that act as important post-transcriptional regulators of gene expression by base-pairing to their target mRNAs, thereby mediating mRNA cleavage or translational repression. Recent data suggest that miRNAs are aberrantly expressed in many human cancers and that they may play significant roles as oncogenes or tumour suppressors. Apart from cancer, miRNAs have also been implicated in viral infections, heart disease and neurological disorders. Hence, disease-associated miRNAs represent a potential new class of targets for antisense-based therapeutics, which may yield patient benefits unobtainable by other therapeutic approaches.

LNA (locked nucleic acid) is the first true conformational analogue of RNA, in which the furanose ring in the sugar-phosphate backbone is locked in an RNA-like, C3’-endo conformation. This conformational restriction results in unprecedented binding affinity between single-stranded LNA oligonucleotides and their complementary RNA targets and high stability in blood and tissues in vivo. LNA-modified oligonucleotides have proven outstanding in microRNA recognition and detection due to their high specificity and affinity. Here we report that short, unconjugated LNA-antimiR oligonucleotides can be used as potent molecules for sequence-specific antagonism of disease-associated miRNAs in vitro and in vivo in rodents and non-human primates.

MicroRNA manipulation as a therapeutic strategy for cardiovascular disease
William S. Marshall, PhD., President and Chief Executive Officer, miRagen Therapeutics, Boulder, CO, USA

Heart failure remains a major unmet medical need, with more than 20 million patients worldwide and a 50% five year mortality rate. Recent studies have demonstrated the fundamental importance of microRNA perturbations in several forms of cardiovascular disease. miRagen’s founders and scientists combine a strong knowledge of cardiovascular medicine with a deep understanding of microRNA biology and chemistry. Leveraging our core capabilities in microRNA profiling, oligonucleotide medicinal chemistry, high throughput cellular analysis, and transgenic and disease models, we select highly potent modulators that target the most relevant microRNAs in a disease process. By correcting microRNA levels we hope to develop breakthrough therapies that target the fundamental pathways responsible for this progressive, fatal condition.

miRNAs in mammary development and breast cancer
Carlos Caldas MD FACP FRCP FMedSci, Professor of Cancer Medicine, Department of Oncology at the University of Cambridge, Functional Breast Cancer Genomics Laboratory Cancer Research UK, Cambridge Research Institute, Li Ka-Shing Centre, Robinson Way, Cambridge CB2 0RE, UK

miRNAs have now been directly implicated in tumor development acting as either oncogenes or tumor suppressor genes. Several studies recently uncovered the key involvement of miRNAs in breast cancer: 1- miRNAs classify breast cancers into different molecular subtypes suggesting miRNA profiling might supersede mRNA profiling in molecular diagnostics and prognostics; 2- miRNA deregulation in breast cancers occurs at different levels, particularly
miRNA processing, providing opportunities for therapeutic targeting; 3- miRNAs are key regulators of normal and malignant mammary stem cells and breast epithelial stem cell assays could be used to test the mechanism of miRNA-stem cell regulation.

Genetic insights in to the mechanism of RISC function in Mice
Dónal O’Carroll Ph.D., Group Leader, EMBL Monterotondo Mouse Biology Unit, Monterotondo Scalo (RM), Italy

Binding of microRNA (miRNA) to mRNA within the RNA–induced silencing complex (RISC) leads to either translational inhibition or to destruction of the target mRNA. Both of these functions are executed by Argonaute 2 (Ago2). Using hematopoiesis in mice as a model system to study the physiological function of Ago2 in vivo, we found that Ago2 controls early development of lymphoid and erythroid cells. We show that the unique and defining feature of Ago2, the Slicer endonuclease activity, is dispensable for hematopoiesis. Instead, we identified Ago2 as a key regulator of miRNA homeostasis. Deficiency in Ago2 impairs miRNA biogenesis from precursor-miRNAs followed by a reduction in miRNA expression levels. Collectively, our data identify Ago2 as a highly specialized member of the Argonaute family with an essential non-redundant Slicer independent function within the mammalian miRNA pathway. 

MicroRNAs in cancer and hematopoiesis
Cesare Peschle, M.D., Chairman and Professor, IRCCS MultiMedica, Milan & Istituto Superiore di Sanità, Roma, Italy

Our laboratori has recently focused on the expression and regulatory function of microRNAs in normal hematopoiesis (1-3) and solid tumors (4-6) including cancer stem cells (7), as well as in heart hypertrophy (8). The studies on microRNA-based control pathways in hematopoietic lineages and oncogenic mechanisms in prostate cancer, melanoma and pediatric neuroblastoma will be discussed.

References
Labbaye C. et al, Nature Cell Biol, 2008
Fontana L. et al, Nature Cell Biol, 2007
Felli N.et al, PNAS, 2005
Bonci D. et al, Nature Med., 2008
Fontana L. et al., PLoS, 2008
Felicetti et al., Canc. Res., 2008
Ricci-Vitiani L. et al., Nature, 2007
Caré A. et al., Nature Med., 2007

MicroRNA regulation of the immune system
Elena Vigorito, PhD., Group Leader, Babraham Institute, Lab. Of Lymphocyte Signaling & Development, Cambridge, UK

MicroRNAs (miRNAs) represent a new class of regulatory genes important for a variety of cellular processes. In the immune system a particular miRNA, miR-155, is rapidly induced in myeloid and lymphoid cells upon activation. Moreover, it is highly expressed in hematological malignancies and it causes myeloid proliferation and B cell expansion when over-expressed in mice. Unraveling how the expression of miRNAs is regulated, which cellular function they influence and identification of their molecular targets under normal conditions is therefore essential to understand their role in cancer. To define the role of miR-155 in vivo, we have analyzed the phenotype of mice in which miR-155 has been ablated. In this presentation, I will summarize what we have learned on the regulation of the immune system by miR-155.

New High-Throughput Facility Expands miRNA Profiling Opportunity
Graham Speight, PhD., Head of Molecular Biology, Oxford Gene Technology, Oxford, UK

Whilst significant progress has been made in genome wide association studies, the biomarkers currently identified account for a small proportion of the overall genetic risk in the disease studied. The race is on to identify more informative biomarkers. OGT (an Agilent Certified Service Provider) currently processing >19,000 samples as part of the Wellcome Trust Case Control Consortium, has expanded its service offering. Here we outline how OGT has implemented a high quality, rapid turnaround microarray capability – which reduces the impact of technical variability and can now be applied to miRNA analysis. 

 

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