title: Glycoprotein Ibalpha promoter drives megakaryocytic lineage-restricted expression after hematopoietic stem cell transduction using a self-inactivating lentiviral vector.
abstract: Megakaryocytic (MK) lineage is an attractive target for cell/gene therapy approaches, aiming at correcting platelet protein deficiencies. However, MK cells are short-lived cells, and their permanent modification requires modification of hematopoietic stem cells with an integrative vector such as a lentiviral vector. Glycoprotein (Gp) IIb promoter, the most studied among the MK regulatory sequences, is also active in stem cells. To strictly limit transgene expression to the MK lineage after transduction of human CD34(+) hematopoietic cells with a lentiviral vector, we looked for a promoter activated later during MK differentiation. Human cord blood, bone marrow, and peripheral-blood mobilized CD34(+) cells were transduced with a human immunodeficiency virus-derived self-inactivating lentiviral vector encoding the green fluorescent protein (GFP) under the transcriptional control of GpIbalpha, GpIIb, or EF1alpha gene regulatory sequences. Both GpIbalpha and GpIIb promoters restricted GFP expression (analyzed by flow cytometry and immunoelectron microscopy) in MK cells among the maturing progeny of transduced cells. However, only the GpIbalpha promoter was strictly MK-specific, whereas GpIIb promoter was leaky in immature progenitor cells not yet engaged in MK cell lineage differentiation. We thus demonstrate the pertinence of using a 328-base-pair fragment of the human GpIbalpha gene regulatory sequence, in the context of a lentiviral vector, to tightly restrict transgene expression to the MK lineage after transduction of human CD34(+) hematopoietic cells. Disclosure of potential conflicts of interest is found at the end of this article.
authors: Institut Cochin, Department of Hematology, Hôpital de Port-Royal, 123 Bd de Port-Royal, Paris 75014, France.
authors: Cécile
authors: C
authors: Lavenu-Bombled
authors: Brigitte
authors: B
authors: Izac
authors: Faézeh
authors: F
authors: Legrand
authors: Marie
authors: M
authors: Cambot
authors: Agathe
authors: A
authors: Vigier
authors: Jean-Marc
authors: JM
authors: Massé
authors: Anne
authors: A
authors: Dubart-Kupperschmitt
mesh_terms: D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism; D002454:Cell Differentiation / Q000235:genetics; D019070:Cell Lineage / Q000235:genetics; D002478:Cells, Cultured; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D018380:Hematopoietic Stem Cell Transplantation; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D016086:Lentivirus; D008533:Megakaryocytes / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D009928:Organ Specificity; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000235:genetics*; D011401:Promoter Regions, Genetic / Q000502:physiology*; D014161:Transduction, Genetic; D038301:Virus Inactivation
chemical_list: D018952:Antigens, CD34; D019038:Platelet Glycoprotein GPIb-IX Complex