CRISPR/Cas9-Mediated Correction of the Sickle Mutation in Human CD34+ cells

Mol Ther. 2016 Sep;24(9):1561-9. doi: 10.1038/mt.2016.148. Epub 2016 Jul 29.


Targeted genome editing technology can correct the sickle cell disease mutation of the β-globin gene in hematopoietic stem cells. This correction supports production of red blood cells that synthesize normal hemoglobin proteins. Here, we demonstrate that Transcription Activator-Like Effector Nucleases (TALENs) and the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9 nuclease system can target DNA sequences around the sickle-cell mutation in the β-globin gene for site-specific cleavage and facilitate precise correction when a homologous donor template is codelivered. Several pairs of TALENs and multiple CRISPR guide RNAs were evaluated for both on-target and off-target cleavage rates. Delivery of the CRISPR/Cas9 components to CD34+ cells led to over 18% gene modification in vitro. Additionally, we demonstrate the correction of the sickle cell disease mutation in bone marrow derived CD34+ hematopoietic stem and progenitor cells from sickle cell disease patients, leading to the production of wild-type hemoglobin. These results demonstrate correction of the sickle mutation in patient-derived CD34+ cells using CRISPR/Cas9 technology.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anemia, Sickle Cell / genetics*
  • Anemia, Sickle Cell / therapy
  • Base Sequence
  • CRISPR-Cas Systems*
  • Cell Line
  • DNA Cleavage
  • Gene Editing*
  • Gene Targeting
  • Genetic Loci
  • Hematopoietic Stem Cells / metabolism*
  • Humans
  • Mutation*
  • Protein Binding
  • RNA, Guide
  • Targeted Gene Repair*
  • Transcription Activator-Like Effector Nucleases / metabolism
  • beta-Globins / genetics*


  • RNA, Guide
  • beta-Globins
  • Transcription Activator-Like Effector Nucleases