Tuesday, June 10

7:30am – 6:00 pm Registration Open

7:30 am Breakfast Workshop (Sponsorships Available)

8:15 Chairperson’s Remarks
Richard Klinghoffer, Ph.D., Research Fellow, Biology, Rosetta Inpharmatics/Merck & Co., Inc.

8:20 Lentivirus-Mediated RNAi Screens 
Richard Klinghoffer, Ph.D., Research Fellow, Biology, Rosetta Inpharmatics/Merck & Co., Inc.
To expand our capacity to interrogate disease-related gene families and signaling cascades by RNAi screens, we have established methods to produce and screen large collections of shRNA-containing lentiviral vectors. We have now used our methods to efficiently generate a screening compatible library of ~14,000 individually arrayed vectors targeting the human druggable genome (approximately 5,000 genes). Screens designed to identify novel regulators of disease have been performed and will be presented.

8:50 Rapid Creation of RNAi Libraries Using shRNA Microarrays
Michael C. Bassik, Ph.D., Post-doctoral Fellow, Laboratory of Michael T. McManus, Department of Cellular and Molecular Pharmacology and Diabetes Center, Department of Microbiology and Immunology, University of California, San Francisco
RNAi has become a widely used and powerful tool for conducting genetic screens. Lentiviral shRNA delivery is most effective for stable gene disruption in diverse cell types, but currently available libraries are limited in a number of critical ways. To address some of these problems, we have developed a quick and efficient strategy to use microarray synthesis of complex shRNA pools in order to generate highly adaptable RNAi libraries. These libraries are inexpensive to create and maintain, can be easily changed to accommodate new vector design, and are less costly and cumbersome to use than standard arrayed libraries. Most importantly, this new strategy allows for much more comprehensive and effective gene targeting. We show here that we can couple the use of these libraries with sorting flow cytometry to identify and quantitate the potency of shRNAs against a number of targets, providing a significant improvement over current screening methods.

9:20 Rational Design Leads to More Potent RNA Interference Targeting Hepatitis B Virus
Anton P. McCaffrey, Ph.D., Assistant Professor, Department of Internal Medicine, University of Iowa
Previously, we conducted proof-of-principle experiments using RNAi to degrade HBV RNAs in mice, and reduce levels of viral proteins and replicated DNA genomes. Recently we expressed the short hairpin RNA (shRNA), HBVU6#2, in HBV transgenic mice. While this RNAi trigger resulted in substantial HBV knockdown in mice, it also resulted in acute toxicity due to competition with the endogenous microRNA machinery. Using rational approaches based on mechanistic insights, we have designed RNAi triggers that are much more potent than the trigger identified in our original screen.

9:50 Networking Coffee Break, Poster and Exhibit Viewing

Panel Discussion:

Intellectual Property Challenges around RNAi Technologies & Therapies 10:45 Strategies for IP Protection around RNAi Therapeutics  Steven Highlander, Ph.D., Partner, Intellectual Property, Fulbright & Jaworski, LLP RNAi technologies present fairly predictable challenges when it comes to securing patent protection. I will review the basic requirements for patent protection and apply each requirement to real world examples of RNAi inventions at various stages of development. Options and strategies for dealing with current U.S. PTO and EP examination practices will be discussed. 11:05 Due Diligence for RNAi; How Do You Look at the Patent Landscape Rochelle K. Seide, Ph.D., Patent Consultant An analysis of the patent landscape surrounding RNAi technology and products is of particular importance in avoiding patent infringement and in licensing and other deals, such as company and/or product acquisitions. The presentation will provide an overview on how to carry out the analysis, including what to look for and how to avoid pitfalls.

11:20 Questions from the Audience

11:45 Keynote Presentation Small RNAs as Markers, Effectors, and Targets of Genetic Change Andrew Fire, Ph.D., Nobel Laureate and Professor, Departments of Pathology and Genetics, Stanford University School of Medicine The “Small-RNA-ome” of a cell or tissue sample gives us a remarkable window into ongoing processes of gene regulation that have biological, procedural, and clinical consequences. This talk will provide several examples in which we hope that Small-RNA-ome determina-tion combined with other analysis will contribute to understanding of (and control over) critical biological processes.

12:30 pm Luncheon Technology Workshops (Sponsorships Available) or Lunch on Your Own

Tuesday June, 10

12:00 pm Registration Open

2:00 Chairperson’s Remarks
Mark A. Kay M.D., Ph.D., Professor, Departments of Pediatrics and Genetics, Stanford University 

2:05 Systemic RNAi Therapeutics for Treating Infection
Mark A. Kay M.D., Ph.D., Professor, Departments of Pediatrics and Genetics, Stanford University 
Gene transfer vectors expressing shRNAs to target specific tissues have been utilized for treating different diseases. Interestingly, over expression of shRNAs can be toxic and even lethal because it interferes with normal microRNA processing. The rate-limiting steps in mammalian tissues as well as effective strategies to maintain a high therapeutic index will be discussed.

2:35 Delivering RNAi Therapeutics
Muthiah Manoharan Ph.D., Vice President, Drug Discovery, Alnylam Pharmaceuticals, Inc.

3:05 Delivery of Therapeutic RNA Interference into the GI tract
Johannes Fruehauf MD, Ph.D., Vice President, Research, Cequent Pharmaceuticals Inc.
Cequent has developed a system that allows the delivery of therapeutic RNA interference into the gastrointestinal tract through oral application. This method, called Transkingdom RNA interference, (tkRNAi), uses nonpathogenic bacteria that are modified to act as manufacturers and carrier vehicles of interfering RNA against genes of interest. Activity has so far been shown across a wide range of targets. APCmin mice are a genetic model of human colon cancer based on dysregulation of beta-catenin (CTNNB1) and the wnt pathway. They develop multiple polyps in their gastrointestinal tract resulting in decreased life span due to chronic obstruction and bleeding. Blockage of CTNNB1 in the gastrointestinal epithelium, e.g. through therapeutic RNA interference, should result in therapeutic or preventive effects. Here we show, that chronic oral treatment of APCmin mice (n=38) with tkRNAi bacteria resulted in a significant decrease of polyp formation through blockage of the CTNNB1 pathway in the gut. Tumor sizes and numbers were reduced and animals displayed fewer polyps with high grade dysplasia after oral treatment with tkRNAi bacteria conferring silencing against CTNNB1. These findings open the possibility of developing RNAi-based drugs for organs and tissues outside of the areas targeted by currently ongoing clinical trials, including the gastrointestinal tract, genitourinary tract, and the skin.

3:50 Networking Refreshment Break, Poster and Exhibit Viewing

4:30 Construction of phi29 DNA-Packaging Motor for Applications in Nanotechnology, Therapy, Diagnosis, and Drug Delivery
Peixuan Guo, Ph.D., Chair in Biomedical Engineering and Director of NIH Nanomedicine Development Center, University of Cincinnati
Bacterial virus phi29 packaging RNA (pRNA) is an ATP-binding component of the DNA packaging motor. pRNA contains aintermolecular interaction domain and a 5’/3’ helical domain. Its unique feature to form dimer, trimer, hexamer and patterned superstructures via the interaction of two interlocking loops makes it a promising tool in nanomedicine. Replacement or insertion of the 5’/3’helical domain with siRNA, ribozyme and receptor-binding aptamer or other therapeutic molecules does not interferer with the formation of the multimers, making it a novel vehicle for targeted therapy, pathogen detection and drug delivery. The chimeric siRNA/pRNA complex induced apoptosis in specific cancer cells, as tested in both cell culture and in animal trials. Such protein-free nanoparticles as therapeutic reagents would allow repeated treatment for chronic diseases.

5:00 In vivo Imaging of siRNA Delivery and Silencing in Tumors
Anna Moore, Ph.D., Associate Professor, Department of Radiology and Director, Molecular Imaging Laboratory, Massachusetts General Hospital
During the past years, RNAi has become an indispensable research instrument in virtually all fields of medical and biological sciences. Its broad applicability (virtually any gene can be silenced), superior efficiency (100-1000-fold compared to antisense oligonucleotides), and exquisite specificity (single nucleotide) could potentially be used to develop a powerful novel treatment paradigm with global relevance to any disease amenable to manipulation at the level of gene expression. The fast developing field of RNA interference requires monitoring of siRNA delivery to targeted organs and evaluating the efficiency of target gene silencing. Molecular imaging techniques represent a powerful tool for real-time non-invasive monitoring of various events at a near microscopic level and have superior advantages over conventional in vitro and cell culture research techniques in biology. Therefore, molecular imaging approach fits perfectly to fulfill the need to monitor siRNA delivery and provides information in a fast, reproducible and non-invasive manner. This presentation will summarize the existing information on various imaging modalities and their application for siRNA imaging.

5:30 Close of Day