Correction of the sickle cell disease mutation in human hematopoietic stem/progenitor cells.
Sickle cell disease (SCD) is characterized by a single point mutation in the seventh codon of the β-globin gene. Site-specific correction of the sickle mutation in hematopoietic stem cells would allow for permanent production of normal red blood cells. Using zinc-finger nucleases (ZFNs) designed to flank the sickle mutation, we demonstrate efficient targeted cleavage at the β-globin locus with minimal off-target modification. By co-delivering a homologous donor template (either an integrase-defective lentiviral vector or a DNA oligonucleotide), high levels of gene modification were achieved in CD34(+) hematopoietic stem and progenitor cells. Modified cells maintained their ability to engraft NOD/SCID/IL2rγ(null) mice and to produce cells from multiple lineages, although with a reduction in the modification levels relative to the in vitro samples. Importantly, ZFN-driven gene correction in CD34(+) cells from the bone marrow of patients with SCD resulted in the production of wild-type hemoglobin tetramers.
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA;
Megan D
http://orcid.org/0000-0002-5011-9582
MD
Hoban
Sangamo BioSciences Inc., Richmond, CA;
Gregory J
http://orcid.org/0000-0002-8509-4653
GJ
Cost
Sangamo BioSciences Inc., Richmond, CA;
Matthew C
MC
Mendel
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA;
Zulema
http://orcid.org/0000-0001-8305-8184
Z
Romero
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA;
Michael L
http://orcid.org/0000-0003-2441-5836
ML
Kaufman
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA;
Alok V
AV
Joglekar
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA;
Michelle
M
Ho
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA;
Dianne
D
Lumaquin
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA;
David
http://orcid.org/0000-0002-2515-6621
D
Gray
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA;
Georgia R
http://orcid.org/0000-0001-5292-3745
GR
Lill
Molecular Biology Interdepartmental PhD Program, University of California, Los Angeles, Los Angeles, CA;
Aaron R
http://orcid.org/0000-0003-4588-2513
AR
Cooper
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA;
Fabrizia
F
Urbinati
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA;
Shantha
S
Senadheera
Sangamo BioSciences Inc., Richmond, CA;
Allen
A
Zhu
Sangamo BioSciences Inc., Richmond, CA;
Pei-Qi
PQ
Liu
Sangamo BioSciences Inc., Richmond, CA;
David E
DE
Paschon
Sangamo BioSciences Inc., Richmond, CA;
Lei
L
Zhang
Sangamo BioSciences Inc., Richmond, CA;
Edward J
EJ
Rebar
Department of Medical Microbiology, Immunology and Cell Biology, Southern Illinois University School of Medicine, Springfield, IL;
Andrew
A
Wilber
Department of General Internal Medicine and Health Services Research, University of California, Los Angeles, Los Angeles, CA; and.
Xiaoyan
X
Wang
Sangamo BioSciences Inc., Richmond, CA;
Philip D
PD
Gregory
Sangamo BioSciences Inc., Richmond, CA;
Michael C
MC
Holmes
Sangamo BioSciences Inc., Richmond, CA;
Andreas
A
Reik
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA;
Roger P
RP
Hollis
Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, Los Angeles, CA; Eli & Edythe Broad Center of Regenerative Medicine & Stem Cell Research, University of California, Los Angeles, Los Angeles, CA.
Donald B
http://orcid.org/0000-0003-1840-6087
DB
Kohn
D000755:Anemia, Sickle Cell / Q000235:genetics* / Q000473:pathology / Q000628:therapy*; D000818:Animals; D018952:Antigens, CD34 / Q000032:analysis; D001483:Base Sequence; D001854:Bone Marrow Cells / Q000378:metabolism / Q000473:pathology; D002478:Cells, Cultured; D004706:Endodeoxyribonucleases / Q000378:metabolism; D005312:Fetal Blood / Q000637:transplantation; D056426:Genetic Loci; D015316:Genetic Therapy; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D008969:Molecular Sequence Data; D009154:Mutation; D016335:Zinc Fingers; D055544:beta-Globins / Q000235:genetics*
D018952:Antigens, CD34; D055544:beta-Globins; D004706:Endodeoxyribonucleases