title: Production of genetically stable high-titer retroviral vectors that carry a human gamma-globin gene under the control of the alpha-globin locus control region.
abstract: The ability to generate stable high-titer vectors that give rise to high levels of expression of transduced globin genes in erythroid cells is a prerequisite for effective retroviral-mediated globin gene therapy. The human beta-globin gene with its immediate flanking sequences does not contain all the regulatory elements necessary for regulated high-level and position-independent expression in erythroid cells. The regulatory element known as the beta-globin locus control region (BetaLCR) can provide a linked Beta-globin gene with these properties. However, addition of BetaLCR sequences to a retrovirus carrying a beta-globin gene increases its genetic instability. We have developed a new generation of retroviral vectors in which a human gamma-globin gene is placed under the control of the alphaLCR, the major regulatory element of the alpha-globin gene cluster. We demonstrate that these retroviruses are genetically stable in producer cell lines and can be produced at high titers that exceed 5 x 10(6) colony-forming units (CFU)/mL. In addition, we show that the transduced gamma-globin gene can be expressed in the adult erythroid environment of mouse erythroleukemia (MEL) cells at a level comparable to that of a single endogenous Betamaj-globin gene. These retroviruses can also transduce primary murine bone marrow progenitor cells as efficiently as retroviruses that carry the neomycin resistance (neor) gene. This new generation of globin retroviral vectors may prove useful for gene therapy of human beta-globin gene disorders such as sickle cell disease and beta-thalassemia.
authors.affiliation: Department of Medicine, Mount Sinai School of Medicine, New York, NY 10029 USA.
authors.forename: S
authors.initials: S
authors.lastname: Ren
authors.forename: B Y
authors.initials: BY
authors.lastname: Wong
authors.forename: J
authors.initials: J
authors.lastname: Li
authors.forename: X N
authors.initials: XN
authors.lastname: Luo
authors.forename: P M
authors.initials: PM
authors.lastname: Wong
authors.forename: G F
authors.initials: GF
authors.lastname: Atweh
mesh_terms: D000755:Anemia, Sickle Cell / Q000235:genetics / Q000628:therapy; D001854:Bone Marrow Cells; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D015672:Erythroid Precursor Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics*; D005914:Globins / Q000235:genetics*; D006801:Humans; D004915:Leukemia, Erythroblastic, Acute / Q000473:pathology; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid; D012190:Retroviridae / Q000235:genetics*; D014407:Tumor Cells, Cultured; D017086:beta-Thalassemia / Q000235:genetics / Q000628:therapy
chemical_list: D011993:Recombinant Fusion Proteins; D005914:Globins; D004121:Dimethyl Sulfoxide