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Insights into the kinetics of siRNA-mediated gene silencing from live-cell and live-animal bioluminescent imaging

Bartlett, Derek W. and Davis, Mark E. (2006) Insights into the kinetics of siRNA-mediated gene silencing from live-cell and live-animal bioluminescent imaging. Nucleic Acids Research, 34 (1). pp. 322-333. ISSN 0305-1048. doi:10.1093/nar/gkj439.

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Small interfering RNA (siRNA) molecules are potent effectors of post-transcriptional gene silencing. Using noninvasive bioluminescent imaging and a mathematical model of siRNA delivery and function, the effects of target-specific and treatment-specific parameters on siRNA-mediated gene silencing are monitored in cells stably expressing the firefly luciferase protein. In vitro, luciferase protein levels recover to pre-treatment values within <1 week in rapidly dividing cell lines, but take longer than 3 weeks to return to steady-state levels in nondividing fibroblasts. Similar results are observed in vivo, with knockdown lasting ~10 days in subcutaneous tumors in A/J mice and 3–4 weeks in the nondividing hepatocytes of BALB/c mice. These data indicate that dilution due to cell division, and not intracellular siRNA half-life, governs the duration of gene silencing under these conditions. To demonstrate the practical use of the model in treatment design, model calculations are used to predict the dosing schedule required to maintain persistent silencing of target proteins with different half-lives in rapidly dividing or nondividing cells. The approach of bioluminescent imaging combined with mathematical modeling provides useful insights into siRNA function and may help expedite the translation of siRNA into clinically relevant therapeutics for disease treatment and management.

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Davis, Mark E.0000-0001-8294-1477
Additional Information:© The Author 2006. Published by Oxford University Press. Received December 2, 2005. Revised December 23, 2005. Accepted December 23, 2005. The authors are especially grateful to D. Petersen and D. Kohn (Children's Hospital Los Angeles) for performing the lentiviral transductions of the luciferase-expressing cell lines; A. McCaffrey and M. Kay (Stanford University) for donating the luciferase-containing plasmid; and J. Heidel (Calando Pharmaceuticals, Inc.) for performing bioluminescent imaging of the mice used in the HPTV studies looking at hepatocyte-specific luciferase expression. This material is based upon work supported under a National Science Foundation Graduate Research Fellowship. This publication was made possible by Grant Number 1 R01 EB004657-01 from the National Institutes of Health (NIH). Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the NIH. Funding to pay the Open Access publication charges for this article was provided by the California Institute of Technology. Conflict of interest statement. None declared.
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Deposited On:17 Jan 2006
Last Modified:08 Nov 2021 19:10

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