Correction of β‐thalassemia major by gene transfer

Authors: Emanuela Anna Roselli Riccardo Mezzadra Marta Claudia Frittoli Giulietta Maruggi Erika Biral Fulvio Mavilio Fabrizio Mastropietro Antonio Amato Giovanni Tonon Chiara Refaldi Maria Domenica Cappellini Marco Andreani Guido Lucarelli Maria Grazia Roncarolo Sarah Marktel Giuliana Ferrari

Publish Date: 2010/08/04

Volume: 2, Issue: 8, Pages: 315-328

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Abstract

β‐Thalassemia is a common monogenic disorder due to mutations in the β‐globin gene and gene therapy based on autologous transplantation of genetically corrected haematopoietic stem cells HSCs holds the promise to treat patients lacking a compatible bone marrow BM donor We recently showed correction of murine β‐thalassemia by gene transfer in HSCs with the GLOBE lentiviral vector LV expressing a transcriptionally regulated human β‐globin gene Here we report successful correction of thalassemia major in human cells by studying a large cohort of pediatric patients of diverse ethnic origin carriers of different mutations and all candidates to BM transplantation Extensive characterization of BM‐derived CD34+ cells before and following gene transfer shows the achievement of high frequency of transduction restoration of haemoglobin A synthesis rescue from apoptosis and correction of ineffective erythropoiesis The procedure does not significantly affect the differentiating potential and the relative proportion of haematopoietic progenitors Analysis of vector integrations shows preferential targeting of transcriptionally active regions without bias for cancer‐related genes Overall these results provide a solid rationale for a future clinical translationβ‐Thalassemia major or Cooleys anaemia leads to a profound anaemia and to death in the first year of life unless regular transfusions are administered So far allogeneic bone marrow transplantation BMT from HLA‐matched donors is the only curative treatment but it is limited to less than 25 of patients Gene therapy based on autologous transplantation of genetically corrected hematopoietic stem cells HSCs represents a promising alternative for patients lacking a suitable donor We have previously developed a lentivirus vector LV GLOBE coding for the human beta globin gene and showed that transplantation of GLOBE‐transduced HSCs corrects thalassemia in murine models To translate these results into the clinics it is now essential to demonstrate the feasibility safety and therapeutic efficacy of this approach by testing the GLOBE LV in the context of human thalassemic cellsCD34+ cells purified from BM aspirates of thalassemia major patients were transduced with GLOBE and induced to differentiate to the erythroblastic lineage The procedure restored haemoglobin production and erythropoiesis overcoming the β‐thalassemia phenotype We sequenced and mapped the GLOBE integration sites in the human cells to assess the risk of integration in potentially dangerous loci and showed that GLOBE integrates at low copy number into the human genome without preferential targeting of proto‐oncogenes tumor‐supressor or cell cycle related genes

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