Institut for Biomedicin

Rasmus Bak

CRISPR/Cas9 β-globin gene targeting in human haematopoietic stem cells

Publikation: Bidrag til tidsskrift/Konferencebidrag i tidsskrift /Bidrag til avisTidsskriftartikelForskningpeer review

DOI

  • Daniel P Dever, Department of Pediatrics, Stanford University, Stanford, California 94305, USA.
  • ,
  • Rasmus O Bak
  • Andreas Reinisch, Department of Medicine, Division of Hematology, Cancer Institute, and Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, California 94305, USA.
  • ,
  • Joab Camarena, Department of Pediatrics, Stanford University, Stanford, California 94305, USA.
  • ,
  • Gabriel Washington, Department of Pediatrics, Stanford University, Stanford, California 94305, USA.
  • ,
  • Carmencita E Nicolas, Department of Pediatrics, Stanford University, Stanford, California 94305, USA.
  • ,
  • Mara Pavel-Dinu, Department of Pediatrics, Stanford University, Stanford, California 94305, USA.
  • ,
  • Nivi Saxena, Department of Pediatrics, Stanford University, Stanford, California 94305, USA.
  • ,
  • Alec B Wilkens, Department of Pediatrics, Stanford University, Stanford, California 94305, USA.
  • ,
  • Sruthi Mantri, Department of Pediatrics, Stanford University, Stanford, California 94305, USA.
  • ,
  • Nobuko Uchida, Stem Cells, Inc. 7707 Gateway Blvd., Suite 140, Newark, California 94560, USA.
  • ,
  • Ayal Hendel, Department of Pediatrics, Stanford University, Stanford, California 94305, USA.
  • ,
  • Anupama Narla, Division of Hematology/Oncology, Department of Pediatrics, Stanford University School of Medicine, Stanford, California 94035, USA.
  • ,
  • Ravindra Majeti, Department of Medicine, Division of Hematology, Cancer Institute, and Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, California 94305, USA.
  • ,
  • Kenneth I Weinberg, Department of Pediatrics, Stanford University, Stanford, California 94305, USA.
  • ,
  • Matthew H Porteus, Department of Pediatrics, Stanford University, Stanford, California 94305, USA.

The β-haemoglobinopathies, such as sickle cell disease and β-thalassaemia, are caused by mutations in the β-globin (HBB) gene and affect millions of people worldwide. Ex vivo gene correction in patient-derived haematopoietic stem cells followed by autologous transplantation could be used to cure β-haemoglobinopathies. Here we present a CRISPR/Cas9 gene-editing system that combines Cas9 ribonucleoproteins and adeno-associated viral vector delivery of a homologous donor to achieve homologous recombination at the HBB gene in haematopoietic stem cells. Notably, we devise an enrichment model to purify a population of haematopoietic stem and progenitor cells with more than 90% targeted integration. We also show efficient correction of the Glu6Val mutation responsible for sickle cell disease by using patient-derived stem and progenitor cells that, after differentiation into erythrocytes, express adult β-globin (HbA) messenger RNA, which confirms intact transcriptional regulation of edited HBB alleles. Collectively, these preclinical studies outline a CRISPR-based methodology for targeting haematopoietic stem cells by homologous recombination at the HBB locus to advance the development of next-generation therapies for β-haemoglobinopathies.

OriginalsprogEngelsk
TidsskriftNature
Vol/bind539
Nummer7629
Sider (fra-til)384-389
Antal sider6
ISSN0028-0836
DOI
StatusUdgivet - 17 nov. 2016
Eksternt udgivetJa

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