Cytosolic delivery of siRNA by ultra-high affinity dsRNA binding proteins
Protein-based methods of siRNA delivery are capable of uniquely specific targeting, but are limited by technical challenges such as low potency or poor biophysical properties. Here, we engineered a series of ultra-high affinity siRNA binders based on the viral protein p19 and developed them into siR...
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| Format: | Article |
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Oxford University Press (OUP)
2018
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| Online Access: | http://hdl.handle.net/1721.1/114564 https://orcid.org/0000-0002-0882-7761 https://orcid.org/0000-0002-0013-3941 https://orcid.org/0000-0002-0493-2863 https://orcid.org/0000-0003-4255-0492 https://orcid.org/0000-0001-5629-4798 https://orcid.org/0000-0003-2398-5896 |
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| author | Sun, Fangdi Yang, Lucy F. Traxlmayr, Michael W. Yu, Yao Xu, Yingda Yang, Nicole Jie Yeon Kauke, Monique Jacqueline Maass, Katie F Langer, Robert S Anderson, Daniel Griffith Wittrup, Karl Dane |
| author2 | Massachusetts Institute of Technology. Institute for Medical Engineering & Science |
| author_facet | Massachusetts Institute of Technology. Institute for Medical Engineering & Science Sun, Fangdi Yang, Lucy F. Traxlmayr, Michael W. Yu, Yao Xu, Yingda Yang, Nicole Jie Yeon Kauke, Monique Jacqueline Maass, Katie F Langer, Robert S Anderson, Daniel Griffith Wittrup, Karl Dane |
| author_sort | Sun, Fangdi |
| collection | MIT |
| description | Protein-based methods of siRNA delivery are capable of uniquely specific targeting, but are limited by technical challenges such as low potency or poor biophysical properties. Here, we engineered a series of ultra-high affinity siRNA binders based on the viral protein p19 and developed them into siRNA carriers targeted to the epidermal growth factor receptor (EGFR). Combined in trans with a previously described endosome-disrupting agent composed of the pore-forming protein Perfringolysin O (PFO), potent silencing was achieved in vitro with no detectable cytotoxicity. Despite concerns that excessively strong siRNA binding could prevent the discharge of siRNA from its carrier, higher affinity continually led to stronger silencing. We found that this improvement was due to both increased uptake of siRNA into the cell and improved pharmacodynamics inside the cell. Mathematical modeling predicted the existence of an affinity optimum that maximizes silencing, after which siRNA sequestration decreases potency. Our study characterizing the affinity dependence of silencing suggests that siRNA-carrier affinity can significantly affect the intracellular fate of siRNA and may serve as a handle for improving the efficiency of delivery. The two-agent delivery system presented here possesses notable biophysical properties and potency, and provide a platform for the cytosolic delivery of nucleic acids. |
| first_indexed | 2024-09-23T12:40:58Z |
| format | Article |
| id | mit-1721.1/114564 |
| institution | Massachusetts Institute of Technology |
| last_indexed | 2024-09-23T12:40:58Z |
| publishDate | 2018 |
| publisher | Oxford University Press (OUP) |
| record_format | dspace |
| spelling | mit-1721.1/1145642022-10-01T10:28:52Z Cytosolic delivery of siRNA by ultra-high affinity dsRNA binding proteins Sun, Fangdi Yang, Lucy F. Traxlmayr, Michael W. Yu, Yao Xu, Yingda Yang, Nicole Jie Yeon Kauke, Monique Jacqueline Maass, Katie F Langer, Robert S Anderson, Daniel Griffith Wittrup, Karl Dane Massachusetts Institute of Technology. Institute for Medical Engineering & Science Massachusetts Institute of Technology. Department of Biological Engineering Massachusetts Institute of Technology. Department of Chemical Engineering Koch Institute for Integrative Cancer Research at MIT Yang, Nicole Jie Yeon Kauke, Monique Jacqueline Maass, Katie F Langer, Robert S Anderson, Daniel Griffith Wittrup, Karl Dane Protein-based methods of siRNA delivery are capable of uniquely specific targeting, but are limited by technical challenges such as low potency or poor biophysical properties. Here, we engineered a series of ultra-high affinity siRNA binders based on the viral protein p19 and developed them into siRNA carriers targeted to the epidermal growth factor receptor (EGFR). Combined in trans with a previously described endosome-disrupting agent composed of the pore-forming protein Perfringolysin O (PFO), potent silencing was achieved in vitro with no detectable cytotoxicity. Despite concerns that excessively strong siRNA binding could prevent the discharge of siRNA from its carrier, higher affinity continually led to stronger silencing. We found that this improvement was due to both increased uptake of siRNA into the cell and improved pharmacodynamics inside the cell. Mathematical modeling predicted the existence of an affinity optimum that maximizes silencing, after which siRNA sequestration decreases potency. Our study characterizing the affinity dependence of silencing suggests that siRNA-carrier affinity can significantly affect the intracellular fate of siRNA and may serve as a handle for improving the efficiency of delivery. The two-agent delivery system presented here possesses notable biophysical properties and potency, and provide a platform for the cytosolic delivery of nucleic acids. National Institutes of Health (U.S.) (Grant CA101830) 2018-04-05T14:48:10Z 2018-04-05T14:48:10Z 2017-06 2017-06 2018-02-23T20:13:14Z Article http://purl.org/eprint/type/JournalArticle 0305-1048 1362-4962 http://hdl.handle.net/1721.1/114564 Yang, Nicole J. et al. “Cytosolic Delivery of siRNA by Ultra-High Affinity dsRNA Binding Proteins.” Nucleic Acids Research 45, 13 (June 2017): 7602–7614 © 2017 The Author(s) https://orcid.org/0000-0002-0882-7761 https://orcid.org/0000-0002-0013-3941 https://orcid.org/0000-0002-0493-2863 https://orcid.org/0000-0003-4255-0492 https://orcid.org/0000-0001-5629-4798 https://orcid.org/0000-0003-2398-5896 http://dx.doi.org/10.1093/NAR/GKX546 Nucleic Acids Research Attribution 4.0 International (CC BY 4.0) https://creativecommons.org/licenses/by/4.0/ application/pdf Oxford University Press (OUP) Nucleic Acids Research |
| spellingShingle | Sun, Fangdi Yang, Lucy F. Traxlmayr, Michael W. Yu, Yao Xu, Yingda Yang, Nicole Jie Yeon Kauke, Monique Jacqueline Maass, Katie F Langer, Robert S Anderson, Daniel Griffith Wittrup, Karl Dane Cytosolic delivery of siRNA by ultra-high affinity dsRNA binding proteins |
| title | Cytosolic delivery of siRNA by ultra-high affinity dsRNA binding proteins |
| title_full | Cytosolic delivery of siRNA by ultra-high affinity dsRNA binding proteins |
| title_fullStr | Cytosolic delivery of siRNA by ultra-high affinity dsRNA binding proteins |
| title_full_unstemmed | Cytosolic delivery of siRNA by ultra-high affinity dsRNA binding proteins |
| title_short | Cytosolic delivery of siRNA by ultra-high affinity dsRNA binding proteins |
| title_sort | cytosolic delivery of sirna by ultra high affinity dsrna binding proteins |
| url | http://hdl.handle.net/1721.1/114564 https://orcid.org/0000-0002-0882-7761 https://orcid.org/0000-0002-0013-3941 https://orcid.org/0000-0002-0493-2863 https://orcid.org/0000-0003-4255-0492 https://orcid.org/0000-0001-5629-4798 https://orcid.org/0000-0003-2398-5896 |
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