title: CRISPR-Cas9 gene repair of hematopoietic stem cells from patients with X-linked chronic granulomatous disease.
abstract: Gene repair of CD34+hematopoietic stem and progenitor cells (HSPCs) may avoid problems associated with gene therapy, such as vector-related mutagenesis and dysregulated transgene expression. We used CRISPR (clustered regularly interspaced short palindromic repeat)/Cas9 (CRISPR-associated 9) to repair a mutation in the CYBB gene of CD34+HSPCs from patients with the immunodeficiency disorder X-linked chronic granulomatous disease (X-CGD). Sequence-confirmed repair of >20% of HSPCs from X-CGD patients restored the function of NADPH (nicotinamide adenine dinucleotide phosphate) oxidase and superoxide radical production in myeloid cells differentiated from these progenitor cells in vitro. Transplant of gene-repaired X-CGD HSPCs into NOD (nonobese diabetic) SCID (severe combined immunodeficient) γc-/-mice resulted in efficient engraftment and production of functional mature human myeloid and lymphoid cells for up to 5 months. Whole-exome sequencing detected no indels outside of the CYBB gene after gene correction. CRISPR-mediated gene editing of HSPCs may be applicable to other CGD mutations and other monogenic disorders of the hematopoietic system.
authors.affiliation: Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA. sderavin@niaid.nih.gov hmalech@nih.gov.
authors.forename: Suk See
authors.initials: SS
authors.lastname: De Ravin
authors.affiliation: MaxCyte Inc., Gaithersburg, MD 20878, USA.
authors.forename: Linhong
authors.initials: L
authors.lastname: Li
authors.affiliation: Cancer Research Technology Program, Leidos Biomedical Research Inc., Frederick, MD 21701, USA.
authors.forename: Xiaolin
authors.initials: X
authors.lastname: Wu
authors.affiliation: Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Uimook
authors.initials: U
authors.lastname: Choi
authors.affiliation: MaxCyte Inc., Gaithersburg, MD 20878, USA.
authors.forename: Cornell
authors.initials: C
authors.lastname: Allen
authors.affiliation: Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Sherry
authors.initials: S
authors.lastname: Koontz
authors.affiliation: Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Janet
authors.initials: J
authors.lastname: Lee
authors.affiliation: Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Narda
authors.initials: N
authors.lastname: Theobald-Whiting
authors.affiliation: Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Jessica
authors.initials: J
authors.lastname: Chu
authors.affiliation: Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Mary
authors.initials: M
authors.lastname: Garofalo
authors.affiliation: Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Colin
authors.initials: C
authors.lastname: Sweeney
authors.affiliation: Laboratory of Immunoregulation, National Institutes of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Lela
authors.initials: L
authors.lastname: Kardava
authors.affiliation: Laboratory of Immunoregulation, National Institutes of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Susan
authors.initials: S
authors.lastname: Moir
authors.affiliation: MaxCyte Inc., Gaithersburg, MD 20878, USA.
authors.forename: Angelia
authors.initials: A
authors.lastname: Viley
authors.affiliation: MaxCyte Inc., Gaithersburg, MD 20878, USA.
authors.forename: Pachai
authors.initials: P
authors.lastname: Natarajan
authors.affiliation: Cancer Research Technology Program, Leidos Biomedical Research Inc., Frederick, MD 21701, USA.
authors.forename: Ling
authors.initials: L
authors.lastname: Su
authors.affiliation: Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Douglas
authors.initials: D
authors.lastname: Kuhns
authors.affiliation: Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Kol A
authors.initials: KA
authors.lastname: Zarember
authors.affiliation: MaxCyte Inc., Gaithersburg, MD 20878, USA.
authors.forename: Madhusudan V
authors.initials: MV
authors.lastname: Peshwa
authors.affiliation: Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA. sderavin@niaid.nih.gov hmalech@nih.gov.
authors.forename: Harry L
authors.initials: HL
authors.lastname: Malech
mesh_terms: D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism; D064113:CRISPR-Cas Systems; D002454:Cell Differentiation; D004260:DNA Repair; D005260:Female; D015316:Genetic Therapy; D006105:Granulomatous Disease, Chronic / Q000235:genetics / Q000628:therapy*; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D016296:Mutagenesis; D009154:Mutation; D000074662:NADPH Oxidase 2 / Q000235:genetics; D009841:Oligonucleotides / Q000235:genetics
chemical_list: D018952:Antigens, CD34; D009841:Oligonucleotides; C056558:CYBB protein, human; D000074662:NADPH Oxidase 2