Skip to main content

Lund University Publications

LUND UNIVERSITY LIBRARIES

Efficient Ablation of Genes in Human Hematopoietic Stem and Effector Cells using CRISPR/Cas9.

Mandal, Pankaj K ; Ferreira, Leonardo M R ; Collins, Ryan ; Meissner, Torsten B ; Boutwell, Christian L ; Friesen, Max ; Vrbanac, Vladimir ; Garrison, Brian S ; Stortchevoi, Alexei and Bryder, David LU , et al. (2014) In Cell Stem Cell 15(5). p.643-652
Abstract
Genome editing via CRISPR/Cas9 has rapidly become the tool of choice by virtue of its efficacy and ease of use. However, CRISPR/Cas9-mediated genome editing in clinically relevant human somatic cells remains untested. Here, we report CRISPR/Cas9 targeting of two clinically relevant genes, B2M and CCR5, in primary human CD4(+) T cells and CD34(+) hematopoietic stem and progenitor cells (HSPCs). Use of single RNA guides led to highly efficient mutagenesis in HSPCs but not in T cells. A dual guide approach improved gene deletion efficacy in both cell types. HSPCs that had undergone genome editing with CRISPR/Cas9 retained multilineage potential. We examined predicted on- and off-target mutations via target capture sequencing in HSPCs and... (More)
Genome editing via CRISPR/Cas9 has rapidly become the tool of choice by virtue of its efficacy and ease of use. However, CRISPR/Cas9-mediated genome editing in clinically relevant human somatic cells remains untested. Here, we report CRISPR/Cas9 targeting of two clinically relevant genes, B2M and CCR5, in primary human CD4(+) T cells and CD34(+) hematopoietic stem and progenitor cells (HSPCs). Use of single RNA guides led to highly efficient mutagenesis in HSPCs but not in T cells. A dual guide approach improved gene deletion efficacy in both cell types. HSPCs that had undergone genome editing with CRISPR/Cas9 retained multilineage potential. We examined predicted on- and off-target mutations via target capture sequencing in HSPCs and observed low levels of off-target mutagenesis at only one site. These results demonstrate that CRISPR/Cas9 can efficiently ablate genes in HSPCs with minimal off-target mutagenesis, which could have broad applicability for hematopoietic cell-based therapy. (Less)
Please use this url to cite or link to this publication:
author
; ; ; ; ; ; ; ; and , et al. (More)
; ; ; ; ; ; ; ; ; ; ; ; ; ; ; and (Less)
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Cell Stem Cell
volume
15
issue
5
pages
643 - 652
publisher
Cell Press
external identifiers
  • pmid:25517468
  • wos:000345012700016
  • scopus:84922671463
ISSN
1934-5909
DOI
10.1016/j.stem.2014.10.004
language
English
LU publication?
yes
additional info
The information about affiliations in this record was updated in December 2015. The record was previously connected to the following departments: Stem Cell Aging (013212073)
id
cb0c1060-2f23-4ba2-bc1f-b88cba5208a9 (old id 4908074)
alternative location
http://www.ncbi.nlm.nih.gov/pubmed/25517468?dopt=Abstract
date added to LUP
2016-04-01 10:36:53
date last changed
2022-04-27 23:32:06
@article{cb0c1060-2f23-4ba2-bc1f-b88cba5208a9,
  abstract     = {{Genome editing via CRISPR/Cas9 has rapidly become the tool of choice by virtue of its efficacy and ease of use. However, CRISPR/Cas9-mediated genome editing in clinically relevant human somatic cells remains untested. Here, we report CRISPR/Cas9 targeting of two clinically relevant genes, B2M and CCR5, in primary human CD4(+) T cells and CD34(+) hematopoietic stem and progenitor cells (HSPCs). Use of single RNA guides led to highly efficient mutagenesis in HSPCs but not in T cells. A dual guide approach improved gene deletion efficacy in both cell types. HSPCs that had undergone genome editing with CRISPR/Cas9 retained multilineage potential. We examined predicted on- and off-target mutations via target capture sequencing in HSPCs and observed low levels of off-target mutagenesis at only one site. These results demonstrate that CRISPR/Cas9 can efficiently ablate genes in HSPCs with minimal off-target mutagenesis, which could have broad applicability for hematopoietic cell-based therapy.}},
  author       = {{Mandal, Pankaj K and Ferreira, Leonardo M R and Collins, Ryan and Meissner, Torsten B and Boutwell, Christian L and Friesen, Max and Vrbanac, Vladimir and Garrison, Brian S and Stortchevoi, Alexei and Bryder, David and Musunuru, Kiran and Brand, Harrison and Tager, Andrew M and Allen, Todd M and Talkowski, Michael E and Rossi, Derrick J and Cowan, Chad A}},
  issn         = {{1934-5909}},
  language     = {{eng}},
  number       = {{5}},
  pages        = {{643--652}},
  publisher    = {{Cell Press}},
  series       = {{Cell Stem Cell}},
  title        = {{Efficient Ablation of Genes in Human Hematopoietic Stem and Effector Cells using CRISPR/Cas9.}},
  url          = {{http://dx.doi.org/10.1016/j.stem.2014.10.004}},
  doi          = {{10.1016/j.stem.2014.10.004}},
  volume       = {{15}},
  year         = {{2014}},
}