Seventh International

RNAi, MicroRNAs -2009-Boston Meeting

on

RNA interference & MicroRNAs: Biochemistry to Drugs & Therapeutics

April 1 - 2, 2009

Venue: Hilton Garden Inn, 420 Totten Pond Road, Waltham, MA, 02451 USA

The Most Popular and The Best Global Event in the RNAi Field!

This Year RNAi Meeting Will Be in Conjunction with Oligonuleotide-2009 Meeting. Two Tracks, One Venue

AGENDA/SPEAKERS   (click here for detailed Agenda)

Click Here to Download the Brochure

Wednesday, April 1, 2009
7:00 – 8:30 A.M: Registration Open
7:30 – 8:45 A.M: Continental breakfast
8:00 - 10:00 A.M: Technology Session I
10.00 AM – 6.00 PM Scientific Sessions

Thursday April 2, 2009
7:00 – 8:30 A.M: Registration Open
7:30 – 8:45 AM: Continental breakfast

8:00 - 10:00 A.M: Technology Session II
10.00 AM – 6.00 PM Scientific Sessions

Agenda will be listed later, please visit again.

Scientific Committee:

	Krishnarao Appasani, PhD., MBA (Chair) Founder & CEO GeneExpression Systems, Inc Waltham, MA USA
	Michelle Lyles, PhD. Vice President, Marketing and Sales Febit, inc. Lexington, MA, USA
	Roberto Weinmann, PhD. Director of Oncology Discovery Bristol Myers Squibb Princeton, NJ, USA
	Dmitry Samarsky, PhD. VP, Technology Development RXi Pharmaceuticals 60 Prescott Street Worcester, MA 01605

	Paul C. Zamecnik, MD. Inaugural Speaker on April 1 1996 Albert Lasker Award Winner Emeritus Professor, Harvard Medical School Massachusetts General Hospital Boston, MA, USA Title: TBA
	Norbert Perrimon, PhD. Keynote Speaker on April 1 Professor of Genetics & HHMI investigator Harvard Medical School Boston, MA, USA Title: High-throughput, genome-wide RNA interference (RNAi) screens
	H. Robert Horvitz, PhD. Keynote Lecture on April 2 Nobel Laureate 2002 (Physiology or Medicine) David H. Koch Professor of Biology & Investigator, Howard Hughes Medical Institute Massachusetts Institute of Technology Cambridge, MA, USA Title: TBA
	Tod Woolf, PhD. Industry Keynote Speaker President & CEO RXi Pharmaceuticals Worcester, MA, USA Title: TBA

Other Speakers:

	David Root, PhD. Broad Institute of MIT & Harvard Cambridge, MA, USA
	Greg Goodall, PhD. Associate Professor of Medicine & Head, Cytokine Research Laboratory Hanson Institute & University of Adelaide Adelaide SA, Australia Title: Regulation of EMT by the miR-200 family of microRNAs
	Akiko Hata, Ph.D. Associate Professor of Biochemistry Tufts University Medical School Boston, MA, USA Title: TBA
	Ye Ding, PhD. Research Scientist Wadsworth Center, Albany, NY, USA Title: TBA
	Andrzej "Andre" Pietrzykowski M.D., Ph.D. Research Assistant Professor of Psychiatry Brudnick Neuropsychiatric Research Institute University of Massachusetts Medical School Worcester, MA 01604, USA Title: microRNA in channel transport and alcohol tolerance
	Dr. Georg Sczakiel Professor Institut für Molekulare Medizin Universität zu Lübeck Lübeck, Germany Title: TBA
	Jingfang Ju, Ph.D. Associate Professor of Pathology Stony Brook University Medical Center Stony Brook, NY, USA Title: Impact of miRNAs in colorectal cancer and osteosarcoma
	Francois Vigneault Ph.D. Post-doctoral Fellow of Genetics Harvard Medical School Boston, MA, USA Title: TBA
	William Marshall, PhD. President & CEO miRagen Therapeutics, Inc. Boulder, CO, USA Title: TBA
	Michelle Lyles, Ph.D. Vice President, Marketing and Sales febit, inc. Lexington, MA, USA  Title: TBA
	Iris Eisenberg – Loebl, PhD. Research Associate at HHMI Children's Hospital Boston Harvard Medical School Boston, MA, USA Title: TBA
	Dmitry Samarsky, PhD. Vice President Technology Development RXi Pharmaceuticals, Inc. Worcester, MA, USA Title: TBA
	Patrick Y. Lu, Ph.D. President and CEO Sirnaomics, Inc. Gaithersburg, MD, USA Title: TBA
	Steven Haney, PhD. Sr. Principal Scientist Pfizer Research Technology Center Cambridge, MA, USA Title: TBA
	Bob D. Brown, Ph.D. Senior Vice President of Research Dicerna Pharmaceuticals, Inc. Watertown, MA, USA Title: TBA
	Paul Anderson, MD., PhD. Associate Chief of Rheumatology Brigham and Women's Hospital Harvard Medical School Boston, MA 02115, USA Title: TBA
	Zachary Zimmerman, PhD. Director of External Alliances Alnylam Pharmaceuticals Cambridge, MA 02142, USA Title: TBA
	Kevin Fettes Associate Director of Process Development Avecia Biotech Milford, MA, USA Title: TBA
	Rene Alvarez, PhD. Principal Scientist Alnylam Pharmaceuticals Cambridge, MA 02142, USA Title: TBA

Key Sessions:

The most-up-to-date developments will be addressed:
The Genetics of RNAi and microRNAs
RNAi-their Molecular Mechanisms in various organisms
RNAi delivery in vivo, in vitro
microRNAs and their diverse roles in biology & disease
Computational and Disease Biology of miRNAs
Drug Target identification and Validation
High throughput genome-wide screenings
Innovative strategies to develop anti-viral, ocular and cancer therapeutics

Exhibitors are welcome to reserve their booth space early!

Please contact if you are interested in speaking in the scientific or Technology workshops of this meeting.

GeneExpression Systems, Inc.
P.O. Box 540170
Waltham, MA 02454 USA
Tel: (781) 891-8181
Fax: (781) 891-8234
E-mail: Genexpsys@expressgenes.com
www.expressgenes.com

Poster Abstract Submission by February 1, 2009

ALL ABSTRACTS

30 Years of Targeting RNA:  A Retrospective
Tod Woolf, Ph.D., President & CEO, RXi Pharmaceuticals, Inc., Worcester, MA, USA

Thirty years ago Paul Zamecnik treated mammalian cells with antisense oligonucleotides.  Since this seminal work, various mechanisms for blocking RNA function in cells have been employed (blocking, RNase H cleavage, ribozymes cleavage and RISC-mediated cleavage) each with practical advantages and disadvantages.  A vast array of modified oligonucleotides chemistries have been explored, but a surprisingly few modified chemistries have gained general use.  This core group of modified chemistries (2’ modified, phosphorothioate, neutral backbones) can be used in a great variety of configurations to impart favorable characteristics on therapeutic oligonucleotides.  The affect of mechanism and chemistry on specificity, toxicity and potency on RNA targeting oligonucleotides will be discussed.

Regulation of EMT by the miR-200 family of microRNAs
Greg Goodall,PhD., Associate Professor &Head, Cytokine Research LaboratoryHanson Institute, & Affiliate Associate Professor of Medicine, University of Adelaide, Adelaide, Australia

Epithelial to mesenchymal transition is proposed to be a key step in the metastasis of epithelial-derived tumours. We found that the miR-200 family plays a dominant role in controlling epithelial phenotype and EMT in vitro through their regulation of the E-cadherin transcriptional repressors ZEB1 and SIP1. Expression of these microRNAs is lost in invasive breast cancer-derived cell lines and primary breast tumour regions that have mesenchymal features. The miR-200 promoter is repressed in mesenchymal cells by ZEB1 and SIP1, establishing a double-negative feedback loop controlling ZEB1/SIP1 and miR-200 family expression that regulates cellular phenotypeand has relevance to the role of these factors in tumor progression.

Technology development for low cost and high-throughput sequencing of microRNAs
Francois Vigneault Ph.D., Post-doctoral Fellow in the Lab of Dr. Church Laboratory
Harvard Medical School,Boston, MA, USA

Francois Vigneault, A. Michael Sismour & George M Church, Harvard Medical School, Dept. of Genetics, Boston, MA, USA
Although most miRNAs studies rely on the detection of previously reported miRNAs, the most powerful approach to identify and quantify expression of new miRNAs remains direct cloning and sequencing. To this extent, we have developed a procedure to facilitate miRNA capture by ligation by barcoding samples, thus allowing multiplexing and considerably minimizing the per-sample cost on next-generation sequencing platforms. With this approach we are planning large scale analysis of the role of miRNAs in population genetics, in the regulation of the VDJ-ome as well as the development of tools for the study of non-coding SNPs on allele specific expression of Pri-miRNAs.

MPEA a miRNA profiling method using on-chip signal amplification; ideal for small sample amounts
Michelle Lyles, PhD., Vice President of Marketing and Sales, febit, inc., Lexington, MA, USA

Small noncoding RNAs (sncRNAs) have moved from a curiosity to a key albeit poorly understood component of gene regulation. Next Generation Sequencing has been at the forefront in the discovery of many such noncoding molecules, yet flexible tools translating this sequence information into affordable high throughput assays are lacking. Here we describe a microfluidic primer extension assay (MPEA) for the detection of sncRNAs employing highly flexible microfluidic microarrays.

MPEA is unique in its integration of conventional hybridization with on-chip enzymatic elongation (primer extension). sncRNAs can be detected in little as 20 ng of total RNA. Importantly, MPEA does not require labeling prior to hybridization and – due to its high sensitivity –amplification is not required. Thus, no bias is introduced by such processes. Here, we present detection of miRNA, and the flexibility of the technology platform enables analysis of any sncRNA, such as piRNAs.

Impact of target secondary structure on function of small RNAs
Ye Ding, PhD., Research Scientist, Developmental Genetics and Bioinformatics
Wadsworth Center, Albany, New York, USA

Target secondary structure predicted by Sfold was found to have a major impact on the efficacy of siRNAs or target recognition by microRNAs. By analyzing a large immunoprecipation (IP) data for worm targets, we identified five significant sequence and target structure features for microRNA targeting. Based on these features, we developed a logistic regression model for genome-scale target prediction. For performance
evaluation using experimental data for C. elegans, our method was found to outperform existing target prediction programs. For Drosophila melanogaster, the method has a high predictive accuracy. Target predictions are available through a database and StarMir web server (http://sfold.wadsworth.org).

Impact of miRNAs in colorectal cancer and osteosarcoma
Jingfang Ju, PhD., Associate Professor, Translational Research Laboratory, Stony Brook University, School of Medicine, Stony Brook, NY, USA

Research involved in the translational regulation of suspected genes in cancer has come to the forefront in recent years. We first reported a regulatory connection between p53 tumor suppressor and certain miRNAs in colon cancer. Some of these miRNAs were associated with clinical outcome of colon cancer treatment. We recently demonstrated that miR-192 and a close related miR-215 can impact cell proliferation and it was directly regulated by p53. Two important targets thymidylate synthase and dihydrofolate reductase of miR-215 were identified in colon cancer and osteosarcoma. We believe these miRNAs will be important as new cancer biomarker and therapeutic targets.

Phosphorothioate-stimulated cellular uptake of naked siRNA indicates new routes for delivery: a quantitative view of siRNA-induced RNAi

Georg Sczakiel, Professor, Universität zu Lübeck, Institut für Molekulare Medizin, Lübeck, Germany

Efficient cellular uptake is a key requirement of the therapeutic and biological application of short silencing RNA (siRNA) in vivo. The phosphorothioate (PS)-stimulated uptake of naked extracellular siRNA into human cells represents a promising alternative as it (i) delivers large amounts of siRNA into cells which is related to measurable though moderate target suppression and (ii) makes use of a caveosomal rather than an endosomal pathway which is used by the majority of known delivery systems. However, even after successful cellular delivery, transport of siRNA to the site of action where it encounters the RNA-induced silencing complex (RISC) and suppresses its target remains to be a major technical hurdle. In many cases siRNA is captured in the endosomal compartment. Conversely, in case of the PS-stimulated mode siRNA accumulates in the perinuclear space which offers new strategies for intracellular functional release of siRNA.

Smad proteins control miRNA maturation 
Akiko Hata, PhD., Associate Professor of Biochemistry, Tufts University School of Medicine, Boston, MA USA

MicroRNAs (miRNAs) are small non-coding RNAs that participate in the spatiotemporal regulation of mRNA and protein synthesis. Aberrant miRNA expression leads to developmental abnormalities and diseases, such as cardiovascular disorders and cancer; however, the stimuli and processes regulating miRNA biogenesis are largely unknown. The transforming growth factor b (TGFb) super-family of growth factors, including bone morphogenetic proteins (BMPs) and TGFbs, orchestrates fundamental biological processes in development and in the homeostasis of adult tissues, including the vasculature. We show that induction of a contractile phenotype in vascular smooth muscle cells (VSMCs) by TGFb and BMPs is mediated by miR-21. Surprisingly, TGFb/BMP signaling promotes a rapid increase in expression of mature miR-21 through a post-transcriptional step, promoting the processing of primary transcripts of miR-21 (pri-miR-21) into precursor miR-21 (pre-miR-21) by the Drosha complex. Smad proteins, the signal transducers of the TGFb family of growth factors, are required for ligand-induced upregulation of pre-miR-21 and mature miR-21. We will discuss a potential mechanism of regulation of miRNA biogenesis by the BMP/TGFb signaling pathway.

MicroRNAs and muscular disorders: New and distinctive players
Iris Eisenberg – Loebl, PhD., Research Associate at HHMI & Children's Hospital Boston
Lou Kunkel’s Lab, Harvard Medical School, Boston, MA, USA

The muscular disorders are a heterogeneous group of inherited diseases characterized by muscle wasting and progressive weakness, resulting in significant morbidity and disability. In recent years, considerable advances have been made in the understanding of the transcriptional and translational mechanisms that regulate skeletal muscle development. Today various signals and transcription factors are known to control the specification and differentiation of skeletal muscle cells. However, the more subtle but decisive processes are still poorly understood. To test whether miRNAs are part of the regulatory circuits in the complexity of skeletal muscle function, a large scale miRNA microarray expression analysis was performed on muscle tissues from a broad spectrum of primary muscle disorders in human, which resulted in the detailed description of the signature pattern of miRNAs associated with ten of the most common muscle conditions. We have reported a comprehensive miRNA expression profile and the identification of new and modifying elements involved in the regulatory networks of muscle. In addition, while five miRNAs were found to be consistently dysregulated in all samples analyzed, suggesting that these miRNAs are involved in a underlying regulatory pathway common among all diseases, other miRNAs were identified to be dysregulated only in one given disease and not in any of the others, thus pointing to their involvement in a unique regulatory mechanism.

Together with the tight post transcriptional regulation at the mRNA level identified in some of the disorders and specific mRNA:miRNA predicted interactions, some of which are directly involved in compensatory secondary response functions and others in muscle regeneration, these findings suggest an important role of miRNAs in the pathology of muscular dystrophy. The subsequent identification of potential target genes, in particular for miRNA 485-5p and miRNA 486, and the unraveling of biological signaling pathways involved in the regulatory level in these disorders, point to an additional dimension of regulation of muscle function mediated by miRNAs.

Rethinking druggable targets in the era of genome-wide association studies and therapeutic RNAi
Steven Haney, Ph.D., Associate Fellow, Biotherapeutics and Bioinovation Center, Pfizer Research Technology Center, Cambridge, MA, USA

RNAi screening for therapeutic targets has historically functioned as a surrogate for screening for small molecule inhibitors, and as such, most screening focused on the major classes of historically druggable targets such as protein kinases and G-protein coupled receptors. However, as oligonucleotide therapeutics make their way to the clinic, RNAi screening can be the direct path to the drug, and screening can focus on new classes of genes that are involved in disease. Genome-wide association studies have identified many clinically relevant genes based on human disease risk, but their specific roles in disease biology are diverse, and present new challenges to the concept of target validation. Technological advances in nucleic acid based therapeutics provide many new opportunities to pursue novel targets based on known human disease risk.

The molecular tale of David and Goliath: How small RNA regulates potassium channel of big conductance
Andrzej Pietrzykowski, M.D., Ph.D., Research Assistant Professor in Psychiatry, University of Massachusetts Medical School, Brudnick Neuropsychiatric Research Institute, Worcester, MA, USA

microRNAs are newly-discovered regulators of gene expression. They are important in a wide variety of cellular processes, with each microRNA regulating expression of hundreds of targets. We will present data showing that alcohol is an important regulator of microRNA expression. This drug of abuse rapidly and specifically upregulates microRNA-9, which in turn, changes mRNA expression of the calcium- and voltage-activated potassium channel of big conductance, called BK. BK channel is abundantly expressed in the brain, and plays an important role in many neuronal functions. Ultimately, alcohol reorganization of BK channel’s mRNA via microRNA-9 leads to an assembly of alcohol-tolerant channels. Moreover, the similar mechanism may contribute to regulation of several other alcohol-relevant genes. These findings highlight an important role of miRNA in the regulation of ion channel gene expression, and open the door to the novel therapeutic strategies in drug addiction.

The power of orthogonal data integration in large scale RNAi studies
Norbert Perrimon, PhD., Professor of Genetics & HHMI investigator Harvard Medical School,
Boston, MA, USA

Signaling networks direct organism development, physiology, and, when altered by mutation, oncogenesis. Characterizing the extent and logic of these networks is essential to understanding specificity in cell fate decisions and mechanisms of oncogenesis and resistance to chemotherapy. Recent studies using cell-based RNA interference (RNAi) screens suggest large numbers of genes regulate signaling pathways.  These functional genomic approaches infer gene contributions to particular cellular outputs but cannot provide network structure directly. Here, we describe a large, integrated network around the canonical receptor tyrosine kinase (RTK)/Ras/extracellular signal-regulated kinases (ERK) signaling pathway by combining parallel RTK- and cell-type-specific genome-wide RNAi screens and tandem affinity purification/mass spectrometry (TAP/MS) protein-protein interaction (PPI) mapping. The integrated network combines results from a total of 2,677 RNAi screen-identified regulators and a PPI network of 710 interactions among 437 proteins. Surprisingly, only a small number of novel proteins beyond the known core pathway score conservatively in both approaches, suggesting that the universal canonical pathway regulating ERK is nearly complete. Despite this, additional experimental and bioinformatic evidence identifies novel and dynamic
subcomplexes, widespread feedback regulation, and biochemically and in vivo validated bona fide interactors and regulators beyond this core pathway likely required in specific contexts. We demonstrate the significance of this more comprehensive RTK/Ras/ERK signaling network derived from orthogonal unbiased approaches by identifying genes mutated in cancers previously linked to isolated canonical pathway components, suggesting a mechanism for these driver mutations and a “pathway-level” susceptibility of these cancers to altered RTK/Ras/ERK signaling. Appreciation of this pathway as a comprehensive network instead of linear cascade will likely be essential to directing appropriate targeted chemotherapies outside of the core pathway and predicting responses to these perturbations.

STP705, A Novel siRNA Therapeutics for Scarless Healing of Skin Wounds
Patrick Y. Lu, Ph.D., President and CEO, Sirnaomics, Inc., Gaithersburg, MD, USA

Vera Simonenko, David Evans, Q. Leng1, James Mixson1 and Patrick Y. Lu, Sirnaomics, Inc. Gaithersburg, MD 20879, USA. 1. Department of Pathology, School of Medicine, University of Maryland

Every year in the United States more than 1.25 million people have burns and 6.5 million have chronic skin ulcers caused by pressure, venous stasis, or diabetes mellitus. The 2006 global market for wound care was estimated to represent about $12 billion and it is expected to grow to $16 billion by 2011. Recent advances in cellular and molecular biology have greatly expanded our understanding of the biologic processes involved in wound repair and tissue regeneration and have led to improvements in wound care. Sirnaomics has developed a novel approach to promote rapid and scarless skin excision wound healing through topical application of a gel like cream containing nanoparticle/siRNA cocktail. The efficient knockdown of target genes in the tissue samples from the skin wounds correlated with a faster closure of skin excision wounds on Balb/c mice. Branched Histidine and Lysine polymer significantly enhanced the siRNA delivery resulted in longer duration and knockdown efficiency of siRNA duplexes. Histological data further demonstrated the therapeutic benefit of the siRNA nanoparticle targeting either single gene or multiple gene targets. A drug candidate STP705 was proposed for IND enabling study to improve scarless healing of various skin excision wounds.