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Bifunctional Short Hairpin RNA (bi-shRNA): Design and Pathway to Clinical Application

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siRNA Design

Part of the book series: Methods in Molecular Biology ((MIMB,volume 942))

Abstract

The discovery of RNA interference (RNAi) engendered great excitement and raised expectations regarding its potential applications in biomedical research and clinical usage. Over the ensuing years, expanded understanding of RNAi and preliminary results from early clinical trials tempered enthusiasm with realistic appraisal resulting in cautious optimism and a better understanding of necessary research and clinical directions. As a result, data from more recent trials are beginning to show encouraging positive clinical outcomes. The capability of delivering a pharmacologically effective dose to the target site while avoiding adverse host reactions still remains a challenge although the delivery technology continues to improve. We have developed a novel vector-driven bifunctional short hairpin RNA (bi-shRNA) technology that harnesses both cleavage-dependent and cleavage-independent RISC loading pathways to enhance knockdown potency. Consequent advantages provided by the bi-shRNA include a lower effective systemic dose than comparator siRNA/shRNA to minimize the potential for off-target side effects, due to its ability to induce both a rapid (inhibition of protein translation) and delayed (mRNA cleavage and degradation) targeting effect depending on protein and mRNA kinetics, and a longer duration of effectiveness for clinical applications. Here, we provide an overview of key molecular methods for the design, construction, quality control, and application of bi-shRNA that we believe will be useful for others interested in utilizing this technology.

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Acknowledgements

The authors gratefully acknowledge the generous support of the W.W. Caruth Jr. Foundation Fund of the Communities Foundation of Texas, the Jasper L. and Jack Denton Wilson Foundation, and the Marilyn Augur Family Foundation. They also wish to thank Susan W. Mill for her assistance in manuscript preparation and submission.

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Authors and Affiliations

  1. Gradalis, Inc., Dallas, TX, USA

    Donald D. Rao, Neil Senzer, Zhaohui Wang, Padmasini Kumar, Chris M. Jay & John Nemunaitis

  2. Mary Crowley Cancer Research Centers, Dallas, TX, USA

    Neil Senzer & John Nemunaitis

  3. Texas Oncology, P.A., Dallas, TX, USA

    Neil Senzer & John Nemunaitis

  4. Medical City Dallas Hospital, Dallas, TX, USA

    Neil Senzer & John Nemunaitis

Authors
  1. Donald D. Rao
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  2. Neil Senzer
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  3. Zhaohui Wang
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  4. Padmasini Kumar
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  5. Chris M. Jay
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  6. John Nemunaitis
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Corresponding author

Correspondence to John Nemunaitis .

Editor information

Editors and Affiliations

  1. , Lineberger Comprehensive Cancer Center, University of North Carolina, West Drive 450, Chapel Hill, 27599, North Carolina, USA

    Debra J. Taxman

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Rao, D.D., Senzer, N., Wang, Z., Kumar, P., Jay, C.M., Nemunaitis, J. (2013). Bifunctional Short Hairpin RNA (bi-shRNA): Design and Pathway to Clinical Application. In: Taxman, D. (eds) siRNA Design. Methods in Molecular Biology, vol 942. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-119-6_14

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  • DOI: https://doi.org/10.1007/978-1-62703-119-6_14

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-118-9

  • Online ISBN: 978-1-62703-119-6

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