title: Therapeutic hemoglobin levels after gene transfer in β-thalassemia mice and in hematopoietic cells of β-thalassemia and sickle cells disease patients.
abstract: Preclinical and clinical studies demonstrate the feasibility of treating β-thalassemia and Sickle Cell Disease (SCD) by lentiviral-mediated transfer of the human β-globin gene. However, previous studies have not addressed whether the ability of lentiviral vectors to increase hemoglobin synthesis might vary in different patients.We generated lentiviral vectors carrying the human β-globin gene with and without an ankyrin insulator and compared their ability to induce hemoglobin synthesis in vitro and in thalassemic mice. We found that insertion of an ankyrin insulator leads to higher, potentially therapeutic levels of human β-globin through a novel mechanism that links the rate of transcription of the transgenic β-globin mRNA during erythroid differentiation with polysomal binding and efficient translation, as reported here for the first time. We also established a preclinical assay to test the ability of this novel vector to synthesize adult hemoglobin in erythroid precursors and in CD34(+) cells isolated from patients affected by β-thalassemia and SCD. Among the thalassemic patients, we identified a subset of specimens in which hemoglobin production can be achieved using fewer copies of the vector integrated than in others. In SCD specimens the treatment with AnkT9W ameliorates erythropoiesis by increasing adult hemoglobin (Hb A) and concurrently reducing the sickling tetramer (Hb S).Our results suggest two major findings. First, we discovered that for the purpose of expressing the β-globin gene the ankyrin element is particularly suitable. Second, our analysis of a large group of specimens from β-thalassemic and SCD patients indicates that clinical trials could benefit from a simple test to predict the relationship between the number of vector copies integrated and the total amount of hemoglobin produced in the erythroid cells of prospective patients. This approach would provide vital information to select the best candidates for these clinical trials, before patients undergo myeloablation and bone marrow transplant.
authors.affiliation: Department of Pediatrics, Division of Hematology-Oncology, Children's Blood and Cancer Foundation Laboratories, Weill Cornell Medical College, New York, New York, United States of America.
authors.forename: Laura
authors.initials: L
authors.lastname: Breda
authors.forename: Carla
authors.initials: C
authors.lastname: Casu
authors.forename: Sara
authors.initials: S
authors.lastname: Gardenghi
authors.forename: Nicoletta
authors.initials: N
authors.lastname: Bianchi
authors.forename: Luca
authors.initials: L
authors.lastname: Cartegni
authors.forename: Mohandas
authors.initials: M
authors.lastname: Narla
authors.forename: Karina
authors.initials: K
authors.lastname: Yazdanbakhsh
authors.forename: Marco
authors.initials: M
authors.lastname: Musso
authors.forename: Deepa
authors.initials: D
authors.lastname: Manwani
authors.forename: Jane
authors.initials: J
authors.lastname: Little
authors.forename: Lawrence B
authors.initials: LB
authors.lastname: Gardner
authors.forename: Dorothy A
authors.initials: DA
authors.lastname: Kleinert
authors.forename: Eugenia
authors.initials: E
authors.lastname: Prus
authors.forename: Eitan
authors.initials: E
authors.lastname: Fibach
authors.forename: Robert W
authors.initials: RW
authors.lastname: Grady
authors.forename: Patricia J
authors.initials: PJ
authors.lastname: Giardina
authors.forename: Roberto
authors.initials: R
authors.lastname: Gambari
authors.forename: Stefano
authors.initials: S
authors.lastname: Rivella
mesh_terms: D000328:Adult; D000755:Anemia, Sickle Cell / Q000097:blood / Q000235:genetics / Q000628:therapy*; D000818:Animals; D017487:Ankyrins / Q000235:genetics; D018952:Antigens, CD34 / Q000378:metabolism; D001483:Base Sequence; D002454:Cell Differentiation / Q000235:genetics; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D015672:Erythroid Precursor Cells / Q000378:metabolism; D015870:Gene Expression; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics; D006454:Hemoglobins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D038101:Insulator Elements / Q000235:genetics; D016086:Lentivirus / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D041681:NIH 3T3 Cells; D055544:beta-Globins / Q000235:genetics; D017086:beta-Thalassemia / Q000097:blood / Q000235:genetics / Q000628:therapy*
chemical_list: D017487:Ankyrins; D018952:Antigens, CD34; D006454:Hemoglobins; D055544:beta-Globins