Journal
NATURE COMMUNICATIONS
Volume 11, Issue 1, Pages -Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/s41467-020-14957-y
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Funding
- Practical Research Project for Rare/Intractable Diseases by AMED [JP19ek0109293]
- T-CiRA Join Research by Takeda Pharmaceutical Company
- Core Center for iPS Cell Research by AMED [JP19bm0104001]
- Acceleration Program for Intractable Diseases Research utilizing disease-specific iPS cells by AMED [JP19bm0804005]
- NCNP
- JSPS [17K15048]
- fellowship program for the Promotion of Internationalization of Research (CiRA, Kyoto University)
- Grants-in-Aid for Scientific Research [17K15048] Funding Source: KAKEN
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Prolonged expression of the CRISPR-Cas9 nuclease and gRNA from viral vectors may cause off-target mutagenesis and immunogenicity. Thus, a transient delivery system is needed for therapeutic genome editing applications. Here, we develop an extracellular nanovesicle-based ribonucleoprotein delivery system named NanoMEDIC by utilizing two distinct homing mechanisms. Chemical induced dimerization recruits Cas9 protein into extracellular nanovesicles, and then a viral RNA packaging signal and two self-cleaving riboswitches tether and release sgRNA into nanovesicles. We demonstrate efficient genome editing in various hard-to-transfect cell types, including human induced pluripotent stem (iPS) cells, neurons, and myoblasts. NanoMEDIC also achieves over 90% exon skipping efficiencies in skeletal muscle cells derived from Duchenne muscular dystrophy (DMD) patient iPS cells. Finally, single intramuscular injection of NanoMEDIC induces permanent genomic exon skipping in a luciferase reporter mouse and in mdx mice, indicating its utility for in vivo genome editing therapy of DMD and beyond.
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