The goal of the work is to advance our original studies on developing the scientific principles required for RNAi-based clinical therapeutics. Our plan is to provide a platform for safe and effective transcription-based RNA interference (RNAi) into vital organs such as the liver. We are specifically interested in developing a therapeutic approach for Hepatitis C Virus Infection (HCV) for which there is still an unmet clinical need. Until recently, research in this area has been hampered by the lack of ability to study replication from an intact viral infection in cell culture and the paucity of animal models. As a result, HCV replicons have been the primary means to study viral replication. We have discovered that some RNAi targets are effective against replicons but relatively inert against bona-fide viral infection that shares identical target sequences. We plan to re-evaluate various regions of the HCV genome for RNAi knockdown based on our new models and information regarding shRNA-vector biology. In addition, we will attempt to definitively evaluate the effectiveness of targeting the HCV minus strand because it is produced in much lower amounts in infected cells. We will determine if targeting both strands simultaneously will provide better protection and lessen the likelihood of quasi-species formation. We plan to use cell culture and humanized mouse liver models with our newly developed recombinant AAV vectors and determine the validity of RNAi as a means to protect against HCV exposure as well as elimination of the virus after establishment of a sustained infection. By the end of the granting period, we believe we will be able to provide important insights into the parameters required for effective vector-based RNAi in vivo and have a robust clinically relevant means of treating HCV infection as a single therapy and/or in combination with other non-nucleic acid based therapeutics.

Public Health Relevance

The goal of this work is to develop the scientific principles and pave the way to develop a safe and efficacious clinical therapy for Hepatitis C virus infection using RNA interference (RNAi). RNAi is a powerful new technology for turning off unwanted genes such as those present in a viral pathogen. The new anti-HCV RNAI therapeutics will be tested for their ability to eliminate and/or prevent HCV infection in newer clinically relevant cell culture and rodent models of HCV infection.

Agency
National Institute of Health (NIH)
Type
Research Project (R01)
Project #
5R01AI071068-09
Application #
8699664
Study Section
Gene and Drug Delivery Systems Study Section (GDD)
Program Officer
Koshy, Rajen
Project Start
Project End
Budget Start
Budget End
Support Year
9
Fiscal Year
2014
Total Cost
Indirect Cost
Name
Stanford University
Department
Pediatrics
Type
Schools of Medicine
DUNS #
City
Stanford
State
CA
Country
United States
Zip Code
94304
Gu, Shuo; Zhang, Yue; Jin, Lan et al. (2014) Weak base pairing in both seed and 3' regions reduces RNAi off-targets and enhances si/shRNA designs. Nucleic Acids Res 42:12169-76
Parameswaran, Poornima; Sklan, Ella; Wilkins, Courtney et al. (2010) Six RNA viruses and forty-one hosts: viral small RNAs and modulation of small RNA repertoires in vertebrate and invertebrate systems. PLoS Pathog 6:e1000764
Grimm, Dirk; Wang, Lora; Lee, Joyce S et al. (2010) Argonaute proteins are key determinants of RNAi efficacy, toxicity, and persistence in the adult mouse liver. J Clin Invest 120:3106-19
Gu, Shuo; Jin, Lan; Zhang, Feijie et al. (2009) Biological basis for restriction of microRNA targets to the 3'untranslated region in mammalian mRNAs. Nat Struct Mol Biol 16:144-50
Haussecker, Dirk; Cao, Dan; Huang, Yong et al. (2008) Capped small RNAs and MOV10 in human hepatitis delta virus replication. Nat Struct Mol Biol 15:714-21
Haussecker, Dirk (2008) The business of RNAi therapeutics. Hum Gene Ther 19:451-62
Streetz, Konrad L; Doyonnas, Regis; Grimm, Dirk et al. (2008) Hepatic parenchymal replacement in mice by transplanted allogeneic hepatocytes is facilitated by bone marrow transplantation and mediated by CD4 cells. Hepatology 47:706-18
Grimm, Dirk; Kay, Mark A (2007) Therapeutic application of RNAi: is mRNA targeting finally ready for prime time? J Clin Invest 117:3633-41
Grimm, Dirk; Kay, Mark A (2007) Combinatorial RNAi: a winning strategy for the race against evolving targets? Mol Ther 15:878-88
Grimm, Dirk; Kay, Mark A (2007) RNAi and gene therapy: a mutual attraction. Hematology Am Soc Hematol Educ Program :473-81