title: High-level expression of hemoglobin A in human thalassemic erythroid progenitor cells following lentiviral vector delivery of an antisense snRNA.
abstract: Mutations at nucleotides 654, 705, or 745 in intron 2 of the human beta-globin gene activate aberrant 3' and 5' splice sites within the intron and prevent correct splicing of beta-globin pre-mRNA, resulting in inhibition of beta-globin synthesis and in consequence beta-thalassemia. Transfection of HeLa cells expressing the 3 thalassemic mutants with modified U7 snRNA (U7.623), containing a sequence antisense to a region between the aberrant splice sites, reduced the incorrect splicing of pre-mRNA and led to increased levels of the correctly spliced beta-globin mRNA and protein. A lentiviral vector carrying the U7.623 gene was effective in restoration of correct splicing in the model cell lines for at least 6 months. Importantly, the therapeutic value of this system was demonstrated in hematopoietic stem cells and erythroid progenitor cells from a patient with IVS2-745/IVS2-1 thalassemia. Twelve days after transduction of the patient cells with the U7.623 lentiviral vector, the levels of correctly spliced beta-globin mRNA and hemoglobin A were approximately 25-fold over background. These results should be regarded as a proof of principle for lentiviral vector-based gene therapy for beta-thalassemia.
authors.affiliation: Curriculum in Genetics and Molecular Biology, University of North Carolina Gene Therapy Center, Department of Pharmacology and Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, 27599, USA.
authors.forename: Marla M
authors.initials: MM
authors.lastname: Vacek
authors.forename: Hong
authors.initials: H
authors.lastname: Ma
authors.forename: Federica
authors.initials: F
authors.lastname: Gemignani
authors.forename: Giuseppina
authors.initials: G
authors.lastname: Lacerra
authors.forename: Tal
authors.initials: T
authors.lastname: Kafri
authors.forename: Ryszard
authors.initials: R
authors.lastname: Kole
mesh_terms: D015672:Erythroid Precursor Cells / Q000378:metabolism*; D015316:Genetic Therapy; D005822:Genetic Vectors; D005914:Globins / Q000235:genetics; D006367:HeLa Cells; D006441:Hemoglobin A / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D009154:Mutation; D012326:RNA Splicing / Q000187:drug effects; D016372:RNA, Antisense / Q000008:administration & dosage / Q000494:pharmacology* / Q000627:therapeutic use; D012342:RNA, Small Nuclear / Q000008:administration & dosage / Q000494:pharmacology* / Q000627:therapeutic use; D013789:Thalassemia / Q000097:blood / Q000235:genetics / Q000628:therapy*; D014162:Transfection
chemical_list: D016372:RNA, Antisense; D012342:RNA, Small Nuclear; C405487:U7 small nuclear RNA; D005914:Globins; D006441:Hemoglobin A