In vivo genome editing via CRISPR/Cas9 mediated homology-independent targeted integration

Handle URI:
http://hdl.handle.net/10754/622418
Title:
In vivo genome editing via CRISPR/Cas9 mediated homology-independent targeted integration
Authors:
Suzuki, Keiichiro; Tsunekawa, Yuji; Hernández-Benítez, Reyna ( 0000-0003-2546-8956 ) ; Wu, Jun; Zhu, Jie; Kim, Euiseok J.; Hatanaka, Fumiyuki; Yamamoto, Mako; Araoka, Toshikazu; Li, Zhe; Kurita, Masakazu; Hishida, Tomoaki; Li, Mo; Aizawa, Emi; Guo, Shicheng; Chen, Song; Goebl, April; Soligalla, Rupa Devi; Qu, Jing; Jiang, Tingshuai; Fu, Xin; Jafari, Maryam; Esteban, Concepcion Rodriguez; Berggren, W. Travis; Lajara, Jeronimo; Nuñez-Delicado, Estrella; Guillen, Pedro; Campistol, Josep M.; Matsuzaki, Fumio; Liu, Guang-Hui; Magistretti, Pierre J. ( 0000-0002-6678-320X ) ; Zhang, Kun; Callaway, Edward M.; Zhang, Kang; Belmonte, Juan Carlos Izpisua
Abstract:
Targeted genome editing via engineered nucleases is an exciting area of biomedical research and holds potential for clinical applications. Despite rapid advances in the field, in vivo targeted transgene integration is still infeasible because current tools are inefficient1, especially for non-dividing cells, which compose most adult tissues. This poses a barrier for uncovering fundamental biological principles and developing treatments for a broad range of genetic disorders2. Based on clustered regularly interspaced short palindromic repeat/Cas9 (CRISPR/Cas9)3, 4 technology, here we devise a homology-independent targeted integration (HITI) strategy, which allows for robust DNA knock-in in both dividing and non-dividing cells in vitro and, more importantly, in vivo (for example, in neurons of postnatal mammals). As a proof of concept of its therapeutic potential, we demonstrate the efficacy of HITI in improving visual function using a rat model of the retinal degeneration condition retinitis pigmentosa. The HITI method presented here establishes new avenues for basic research and targeted gene therapies.
KAUST Department:
4700 King Abdullah University of Science and Technology (KAUST) Thuwal 23955-6900, Saudi Arabia
Citation:
Suzuki K, Tsunekawa Y, Hernandez-Benitez R, Wu J, Zhu J, et al. (2016) In vivo genome editing via CRISPR/Cas9 mediated homology-independent targeted integration. Nature 540: 144–149. Available: http://dx.doi.org/10.1038/nature20565.
Publisher:
Springer Nature
Journal:
Nature
KAUST Grant Number:
OSR-2015-CRG4-2631
Issue Date:
15-Nov-2016
DOI:
10.1038/nature20565
Type:
Article
ISSN:
0028-0836; 1476-4687
Sponsors:
We are grateful to M. Kay, Z. Y. Chen, G. Lemke and P. G. Burrola for sharing experimental materials; J. Naughton, L. Lisowski and J. Marlett for AAV production; C. Fine, J. Olvera, E. O’Connor and K. E. Marquez for cell sorting; D. Okamura and M. Jacobs for mouse surgery and histology processing; D. Skowronska-Krawczyk for rat experiments; N. V. Gohad, T. Whitfield, I. M. Verma, J. Ogawa, T. Hara, U. Manor and J. Santini for imaging; L. Greg, Y. S. Kida and F. Osakada for valuable discussions; D. O’Keefe for proofreading the manuscript and M. Schwarz for administrative help. Core Facilities were utilized at the Salk Institute (support from: NIH-NCI CCSG: P30 014195, NINDS R24NS092943, and NEI P30 EY019005) and UCSD Neuroscience core grant P30 NS047101. R.H.B. was supported by a CONACYT fellowship of Mexico. J.Z. and T.J. were supported by 973 Program (2013CB967504, 2015CB964600) and 863 Program (2014AA021604). T.H. was partially supported by a Nomis Foundation Fellowship. E.J.K. is a Biogen-IDEC Fellow of the Life Science Research Foundation. M.Y. was partially supported by the Salk Women & Science Special Award. X.F. was supported by NSFC (No. 81601872). G.H.L. and J.Q. were supported by the National Basic Research Program of China (973 Program; 2015CB964800, 2014CB910503, 2013CB967504), National Natural Science Foundation of China (81625009, 81371342, 81271266), the National High Technology Research and Development Program of China (2015AA020307, 2014AA021604), and Program of Beijing Municipal Science and Technology Commission (Z151100003915072). F.M. was supported by RIKEN funding for Development and Regeneration. Ku.Z. was supported by NIH grant R01HL123755. P.J.M. and J.C.I.B. were supported by the King Abdullah University of Science and Technology (KAUST) Office of Sponsored Research (OSR) under award no. OSR-2015-CRG4-2631. Work in the laboratory of J.C.I.B. was supported by The Leona M. and Harry B. Helmsley Charitable Trust (2012-PG-MED002), the G. Harold and Leila Y. Mathers Charitable Foundation, NIH (R01HL123755), The McKnight Foundation, The Moxie Foundation, Fundacion Dr. Pedro Guillen and Universidad Católica San Antonio de Murcia (UCAM).
Additional Links:
http://dx.doi.org/10.1038/nature20565
Appears in Collections:
Articles

Full metadata record

DC FieldValue Language
dc.contributor.authorSuzuki, Keiichiroen
dc.contributor.authorTsunekawa, Yujien
dc.contributor.authorHernández-Benítez, Reynaen
dc.contributor.authorWu, Junen
dc.contributor.authorZhu, Jieen
dc.contributor.authorKim, Euiseok J.en
dc.contributor.authorHatanaka, Fumiyukien
dc.contributor.authorYamamoto, Makoen
dc.contributor.authorAraoka, Toshikazuen
dc.contributor.authorLi, Zheen
dc.contributor.authorKurita, Masakazuen
dc.contributor.authorHishida, Tomoakien
dc.contributor.authorLi, Moen
dc.contributor.authorAizawa, Emien
dc.contributor.authorGuo, Shichengen
dc.contributor.authorChen, Songen
dc.contributor.authorGoebl, Aprilen
dc.contributor.authorSoligalla, Rupa Devien
dc.contributor.authorQu, Jingen
dc.contributor.authorJiang, Tingshuaien
dc.contributor.authorFu, Xinen
dc.contributor.authorJafari, Maryamen
dc.contributor.authorEsteban, Concepcion Rodriguezen
dc.contributor.authorBerggren, W. Travisen
dc.contributor.authorLajara, Jeronimoen
dc.contributor.authorNuñez-Delicado, Estrellaen
dc.contributor.authorGuillen, Pedroen
dc.contributor.authorCampistol, Josep M.en
dc.contributor.authorMatsuzaki, Fumioen
dc.contributor.authorLiu, Guang-Huien
dc.contributor.authorMagistretti, Pierre J.en
dc.contributor.authorZhang, Kunen
dc.contributor.authorCallaway, Edward M.en
dc.contributor.authorZhang, Kangen
dc.contributor.authorBelmonte, Juan Carlos Izpisuaen
dc.date.accessioned2017-01-02T09:28:30Z-
dc.date.available2017-01-02T09:28:30Z-
dc.date.issued2016-11-15en
dc.identifier.citationSuzuki K, Tsunekawa Y, Hernandez-Benitez R, Wu J, Zhu J, et al. (2016) In vivo genome editing via CRISPR/Cas9 mediated homology-independent targeted integration. Nature 540: 144–149. Available: http://dx.doi.org/10.1038/nature20565.en
dc.identifier.issn0028-0836en
dc.identifier.issn1476-4687en
dc.identifier.doi10.1038/nature20565en
dc.identifier.urihttp://hdl.handle.net/10754/622418-
dc.description.abstractTargeted genome editing via engineered nucleases is an exciting area of biomedical research and holds potential for clinical applications. Despite rapid advances in the field, in vivo targeted transgene integration is still infeasible because current tools are inefficient1, especially for non-dividing cells, which compose most adult tissues. This poses a barrier for uncovering fundamental biological principles and developing treatments for a broad range of genetic disorders2. Based on clustered regularly interspaced short palindromic repeat/Cas9 (CRISPR/Cas9)3, 4 technology, here we devise a homology-independent targeted integration (HITI) strategy, which allows for robust DNA knock-in in both dividing and non-dividing cells in vitro and, more importantly, in vivo (for example, in neurons of postnatal mammals). As a proof of concept of its therapeutic potential, we demonstrate the efficacy of HITI in improving visual function using a rat model of the retinal degeneration condition retinitis pigmentosa. The HITI method presented here establishes new avenues for basic research and targeted gene therapies.en
dc.description.sponsorshipWe are grateful to M. Kay, Z. Y. Chen, G. Lemke and P. G. Burrola for sharing experimental materials; J. Naughton, L. Lisowski and J. Marlett for AAV production; C. Fine, J. Olvera, E. O’Connor and K. E. Marquez for cell sorting; D. Okamura and M. Jacobs for mouse surgery and histology processing; D. Skowronska-Krawczyk for rat experiments; N. V. Gohad, T. Whitfield, I. M. Verma, J. Ogawa, T. Hara, U. Manor and J. Santini for imaging; L. Greg, Y. S. Kida and F. Osakada for valuable discussions; D. O’Keefe for proofreading the manuscript and M. Schwarz for administrative help. Core Facilities were utilized at the Salk Institute (support from: NIH-NCI CCSG: P30 014195, NINDS R24NS092943, and NEI P30 EY019005) and UCSD Neuroscience core grant P30 NS047101. R.H.B. was supported by a CONACYT fellowship of Mexico. J.Z. and T.J. were supported by 973 Program (2013CB967504, 2015CB964600) and 863 Program (2014AA021604). T.H. was partially supported by a Nomis Foundation Fellowship. E.J.K. is a Biogen-IDEC Fellow of the Life Science Research Foundation. M.Y. was partially supported by the Salk Women & Science Special Award. X.F. was supported by NSFC (No. 81601872). G.H.L. and J.Q. were supported by the National Basic Research Program of China (973 Program; 2015CB964800, 2014CB910503, 2013CB967504), National Natural Science Foundation of China (81625009, 81371342, 81271266), the National High Technology Research and Development Program of China (2015AA020307, 2014AA021604), and Program of Beijing Municipal Science and Technology Commission (Z151100003915072). F.M. was supported by RIKEN funding for Development and Regeneration. Ku.Z. was supported by NIH grant R01HL123755. P.J.M. and J.C.I.B. were supported by the King Abdullah University of Science and Technology (KAUST) Office of Sponsored Research (OSR) under award no. OSR-2015-CRG4-2631. Work in the laboratory of J.C.I.B. was supported by The Leona M. and Harry B. Helmsley Charitable Trust (2012-PG-MED002), the G. Harold and Leila Y. Mathers Charitable Foundation, NIH (R01HL123755), The McKnight Foundation, The Moxie Foundation, Fundacion Dr. Pedro Guillen and Universidad Católica San Antonio de Murcia (UCAM).en
dc.publisherSpringer Natureen
dc.relation.urlhttp://dx.doi.org/10.1038/nature20565en
dc.titleIn vivo genome editing via CRISPR/Cas9 mediated homology-independent targeted integrationen
dc.typeArticleen
dc.contributor.department4700 King Abdullah University of Science and Technology (KAUST) Thuwal 23955-6900, Saudi Arabiaen
dc.identifier.journalNatureen
dc.contributor.institutionGene Expression Laboratory, Salk Institute for Biological Studies, 10010 N. Torrey Pines Rd, La Jolla, California 92037, USAen
dc.contributor.institutionLaboratory for Cell Asymmetry, RIKEN Center for Developmental Biology, 2-2-3 Minatojima-Minamimachi, Chuo-ku, Kobe 650-0047, Japanen
dc.contributor.institutionUniversidad Católica San Antonio de Murcia (UCAM) Campus de los Jerónimos, no. 135 Guadalupe 30107, Murcia, Spainen
dc.contributor.institutionGuangzhou Women and Children’s Medical Center, Guangzhou Medical University, Guangzhou 510623, Chinaen
dc.contributor.institutionShiley Eye Institute, Institute for Genomic Medicine, Institute of Engineering in Medicine, University of California, San Diego, 9500 Gilman Drive #0946, La Jolla, California 92023, USAen
dc.contributor.institutionSystems Neurobiology Laboratory, Salk Institute for Biological Studies, 10010 N. Torrey Pines Rd., La Jolla, California 92037, USAen
dc.contributor.institutionBioengineering, University of California, San Diego, 9500 Gilman Drive, MC0412, La Jolla, California 92093-0412, USAen
dc.contributor.institutionState Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, Chinaen
dc.contributor.institutionUniversity of Chinese Academy of Sciences, Beijing 100049, Chinaen
dc.contributor.institutionGuangzhou EliteHealth Biological Pharmaceutical Technology Company Ltd, Guangzhou 510005, Chinaen
dc.contributor.institutionSalk Institute for Biological Studies, 10010 N. Torrey Pines Rd, La Jolla, California 92037, USAen
dc.contributor.institutionFundación Dr. Pedro Guillen, Investigación Biomedica de Clinica CEMTRO, Avenida Ventisquero de la Condesa, 42, 28035 Madrid, Spainen
dc.contributor.institutionHospital Clinic, University of Barcelona, IDIBAPS, 08036 Barcelona, Spainen
dc.contributor.institutionNational Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, Chinaen
dc.contributor.institutionKey Laboratory of Regenerative Medicine of Ministry of Education, Institute of Aging and Regenerative Medicine, Jinan University, Guangzhou 510632, Chinaen
dc.contributor.institutionBeijing Institute for Brain Disorders, Beijing 100069, Chinaen
dc.contributor.institutionMolecular Medicine Research Center, West China Hospital, Sichuan University, Chengdu 610041, Chinaen
dc.contributor.institutionVeterans Administration Healthcare System, San Diego, California 92093, USAen
kaust.authorHernández-Benítez, Reynaen
kaust.authorMagistretti, Pierre J.en
kaust.grant.numberOSR-2015-CRG4-2631en
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