IDEAS home Printed from https://ideas.repec.org/a/nat/natcom/v14y2023i1d10.1038_s41467-023-42036-5.html
   My bibliography  Save this article

CRISPR-Cas9 engineering of the RAG2 locus via complete coding sequence replacement for therapeutic applications

Author

Listed:
  • Daniel Allen

    (Bar-Ilan University)

  • Orli Knop

    (Bar-Ilan University)

  • Bryan Itkowitz

    (Bar-Ilan University)

  • Nechama Kalter

    (Bar-Ilan University)

  • Michael Rosenberg

    (Bar-Ilan University)

  • Ortal Iancu

    (Bar-Ilan University)

  • Katia Beider

    (The Division of Hematology and Bone Marrow Transplantation, Chaim Sheba Medical Center, Tel-Hashomer)

  • Yu Nee Lee

    (Tel Aviv University
    Edmond and Lily Safra Children’s Hospital, Sheba Medical Center)

  • Arnon Nagler

    (The Division of Hematology and Bone Marrow Transplantation, Chaim Sheba Medical Center, Tel-Hashomer
    Tel Aviv University)

  • Raz Somech

    (Tel Aviv University
    Edmond and Lily Safra Children’s Hospital, Sheba Medical Center)

  • Ayal Hendel

    (Bar-Ilan University)

Abstract

RAG2-SCID is a primary immunodeficiency caused by mutations in Recombination-activating gene 2 (RAG2), a gene intimately involved in the process of lymphocyte maturation and function. ex-vivo manipulation of a patient’s own hematopoietic stem and progenitor cells (HSPCs) using CRISPR-Cas9/rAAV6 gene editing could provide a therapeutic alternative to the only current treatment, allogeneic hematopoietic stem cell transplantation (HSCT). Here we show an innovative RAG2 correction strategy that replaces the entire endogenous coding sequence (CDS) for the purpose of preserving the critical endogenous spatiotemporal gene regulation and locus architecture. Expression of the corrective transgene leads to successful development into CD3+TCRαβ+ and CD3+TCRγδ+ T cells and promotes the establishment of highly diverse TRB and TRG repertoires in an in-vitro T-cell differentiation platform. Thus, our proof-of-concept study holds promise for safer gene therapy techniques of tightly regulated genes.

Suggested Citation

  • Daniel Allen & Orli Knop & Bryan Itkowitz & Nechama Kalter & Michael Rosenberg & Ortal Iancu & Katia Beider & Yu Nee Lee & Arnon Nagler & Raz Somech & Ayal Hendel, 2023. "CRISPR-Cas9 engineering of the RAG2 locus via complete coding sequence replacement for therapeutic applications," Nature Communications, Nature, vol. 14(1), pages 1-16, December.
    • Handle: RePEc:nat:natcom:v:14:y:2023:i:1:d:10.1038_s41467-023-42036-5
    DOI: 10.1038/s41467-023-42036-5
    as

    Download full text from publisher

    File URL: https://www.nature.com/articles/s41467-023-42036-5
    File Function: Abstract
    Download Restriction: no
    File URL: https://libkey.io/10.1038/s41467-023-42036-5?utm_source=ideas
    LibKey link: if access is restricted and if your library uses this service, LibKey will redirect you to where you can use your library subscription to access this item
    ---><---

    References listed on IDEAS

    as
    1. Daniel P. Dever & Rasmus O. Bak & Andreas Reinisch & Joab Camarena & Gabriel Washington & Carmencita E. Nicolas & Mara Pavel-Dinu & Nivi Saxena & Alec B. Wilkens & Sruthi Mantri & Nobuko Uchida & Ayal, 2016. "CRISPR/Cas9 β-globin gene targeting in human haematopoietic stem cells," Nature, Nature, vol. 539(7629), pages 384-389, November.
    2. Mara Pavel-Dinu & Volker Wiebking & Beruh T. Dejene & Waracharee Srifa & Sruthi Mantri & Carmencita E. Nicolas & Ciaran Lee & Gang Bao & Eric J. Kildebeck & Niraj Punjya & Camille Sindhu & Matthew A. , 2019. "Gene correction for SCID-X1 in long-term hematopoietic stem cells," Nature Communications, Nature, vol. 10(1), pages 1-15, December.
    3. Natalia Gomez-Ospina & Samantha G. Scharenberg & Nathalie Mostrel & Rasmus O. Bak & Sruthi Mantri & Rolen M. Quadros & Channabasavaiah B. Gurumurthy & Ciaran Lee & Gang Bao & Carlos J. Suarez & Shauka, 2019. "Human genome-edited hematopoietic stem cells phenotypically correct Mucopolysaccharidosis type I," Nature Communications, Nature, vol. 10(1), pages 1-14, December.
    4. Ido Amit & Ortal Iancu & Alona Levy-Jurgenson & Gavin Kurgan & Matthew S. McNeill & Garrett R. Rettig & Daniel Allen & Dor Breier & Nimrod Ben Haim & Yu Wang & Leon Anavy & Ayal Hendel & Zohar Yakhini, 2021. "CRISPECTOR provides accurate estimation of genome editing translocation and off-target activity from comparative NGS data," Nature Communications, Nature, vol. 12(1), pages 1-11, December.
    Full references (including those not matched with items on IDEAS)

    Most related items

    These are the items that most often cite the same works as this one and are cited by the same works as this one.
    1. Ron Baik & M. Kyle Cromer & Steve E. Glenn & Christopher A. Vakulskas & Kay O. Chmielewski & Amanda M. Dudek & William N. Feist & Julia Klermund & Suzette Shipp & Toni Cathomen & Daniel P. Dever & Mat, 2024. "Transient inhibition of 53BP1 increases the frequency of targeted integration in human hematopoietic stem and progenitor cells," Nature Communications, Nature, vol. 15(1), pages 1-14, December.
    2. Juan A. Perez-Bermejo & Oghene Efagene & William M. Matern & Jeffrey K. Holden & Shaheen Kabir & Glen M. Chew & Gaia Andreoletti & Eniola Catton & Craig L. Ennis & Angelica Garcia & Trevor L. Gerstenb, 2024. "Functional screening in human HSPCs identifies optimized protein-based enhancers of Homology Directed Repair," Nature Communications, Nature, vol. 15(1), pages 1-16, December.
    3. Sandra Wimberger & Nina Akrap & Mike Firth & Johan Brengdahl & Susanna Engberg & Marie K. Schwinn & Michael R. Slater & Anders Lundin & Pei-Pei Hsieh & Songyuan Li & Silvia Cerboni & Jonathan Sumner &, 2023. "Simultaneous inhibition of DNA-PK and Polϴ improves integration efficiency and precision of genome editing," Nature Communications, Nature, vol. 14(1), pages 1-18, December.
    4. M. Kyle Cromer & Valentin V. Barsan & Erich Jaeger & Mengchi Wang & Jessica P. Hampton & Feng Chen & Drew Kennedy & Jenny Xiao & Irina Khrebtukova & Ana Granat & Tiffany Truong & Matthew H. Porteus, 2022. "Ultra-deep sequencing validates safety of CRISPR/Cas9 genome editing in human hematopoietic stem and progenitor cells," Nature Communications, Nature, vol. 13(1), pages 1-11, December.

    More about this item

    Statistics

    Access and download statistics

    Corrections

    All material on this site has been provided by the respective publishers and authors. You can help correct errors and omissions. When requesting a correction, please mention this item's handle: RePEc:nat:natcom:v:14:y:2023:i:1:d:10.1038_s41467-023-42036-5. See general information about how to correct material in RePEc.

    If you have authored this item and are not yet registered with RePEc, we encourage you to do it here. This allows to link your profile to this item. It also allows you to accept potential citations to this item that we are uncertain about.

    If CitEc recognized a bibliographic reference but did not link an item in RePEc to it, you can help with this form .

    If you know of missing items citing this one, you can help us creating those links by adding the relevant references in the same way as above, for each refering item. If you are a registered author of this item, you may also want to check the "citations" tab in your RePEc Author Service profile, as there may be some citations waiting for confirmation.

    For technical questions regarding this item, or to correct its authors, title, abstract, bibliographic or download information, contact: Sonal Shukla or Springer Nature Abstracting and Indexing (email available below). General contact details of provider: http://www.nature.com .

    Please note that corrections may take a couple of weeks to filter through the various RePEc services.

    IDEAS is a RePEc service. RePEc uses bibliographic data supplied by the respective publishers.