title: Targeted disruption of the CCR5 gene in human hematopoietic stem cells stimulated by peptide nucleic acids.
abstract: Peptide nucleic acids (PNAs) bind duplex DNA in a sequence-specific manner, creating triplex structures that can provoke DNA repair and produce genome modification. CCR5 encodes a chemokine receptor required for HIV-1 entry into human cells, and individuals carrying mutations in this gene are resistant to HIV-1 infection. Transfection of human cells with PNAs targeted to the CCR5 gene, plus donor DNAs designed to introduce stop codons mimicking the naturally occurring CCR5-delta32 mutation, produced 2.46% targeted gene modification. CCR5 modification was confirmed at the DNA, RNA, and protein levels and was shown to confer resistance to infection with HIV-1. Targeting of CCR5 was achieved in human CD34(+) hematopoietic stem cells (HSCs) with subsequent engraftment into mice and persistence of the gene modification more than four months posttransplantation. This work suggests a therapeutic strategy for CCR5 knockout in HSCs from HIV-1-infected individuals, rendering cells resistant to HIV-1 and preserving immune system function.
authors.affiliation: Departments of Therapeutic Radiology and Genetics, Yale University School of Medicine, New Haven, CT 06510, USA.
authors.forename: Erica B
authors.initials: EB
authors.lastname: Schleifman
authors.forename: Ranjit
authors.initials: R
authors.lastname: Bindra
authors.forename: Jean
authors.initials: J
authors.lastname: Leif
authors.forename: Jacob
authors.initials: J
authors.lastname: del Campo
authors.forename: Faye A
authors.initials: FA
authors.lastname: Rogers
authors.forename: Pradeep
authors.initials: P
authors.lastname: Uchil
authors.forename: Olaf
authors.initials: O
authors.lastname: Kutsch
authors.forename: Leonard D
authors.initials: LD
authors.lastname: Shultz
authors.forename: Priti
authors.initials: P
authors.lastname: Kumar
authors.forename: Dale L
authors.initials: DL
authors.lastname: Greiner
authors.forename: Peter M
authors.initials: PM
authors.lastname: Glazer
mesh_terms: D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D065100:CCR5 Receptor Antagonists; D002460:Cell Line; D018388:Codon, Terminator; D004260:DNA Repair; D018390:Gene Targeting / Q000379:methods; D015658:HIV Infections / Q000276:immunology / Q000517:prevention & control; D015497:HIV-1 / Q000502:physiology; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000378:metabolism* / Q000821:virology; D006801:Humans; D051379:Mice; D009154:Mutation; D020135:Peptide Nucleic Acids / Q000737:chemistry / Q000494:pharmacology*; D019713:Receptors, CCR5 / Q000235:genetics / Q000378:metabolism*
chemical_list: D018952:Antigens, CD34; D065100:CCR5 Receptor Antagonists; D018388:Codon, Terminator; D020135:Peptide Nucleic Acids; D019713:Receptors, CCR5