title: Mobilized blood CD34+ cells transduced and selected with a clinically applicable protocol reconstitute lymphopoiesis in SCID-Hu mice.
abstract: We developed a clinically applicable gene transfer procedure into mobilized peripheral blood (MPB) CD34(+) hematopoietic progenitor cells, based on single viral exposure and selection of engineered cells. CD34(+) cells were transduced with a retroviral vector carrying the truncated form of the nerve growth factor receptor (Delta NGFR) marker gene, and immunoselected for Delta NGFR expression. Optimal time and procedure for viral exposure, length of culture, and transgene expression of MPB CD34(+) cells were determined using in vitro assays. The multipotent capacity of MPB CD34(+)-transduced cells was demonstrated in the SCID-hu bone/liver/thymus mouse model. Transduced Delta NGFR(+) cells retained 50% of long-term culture-colony forming cells (LTC-CFC) compared to unmanipulated CD34(+) cells. In SCID-hu mice, 52% of CD45(+) cells, 27% of CD34(+) cells, 49% of B cells, and more than 50% of T cells were derived from transplanted CD34(+)/Delta NGFR(+) cells. Furthermore, transplantation of purified transduced cells greatly reduced the competition with untransduced progenitors occurring in unselected grafts. These data demonstrate that MPB CD34(+) cells, transduced with a single viral exposure and selected by transgene expression, retain multilineage reconstitution capacity and remarkable transgene expression.
authors.affiliation: San Raffaele Telethon Institute for Gene Therapy (HSR-TIGET), 20132 Milan, Italy.
authors.forename: Sara
authors.initials: S
authors.lastname: Deola
authors.forename: Samantha
authors.initials: S
authors.lastname: Scaramuzza
authors.forename: Roberto Sciarretta
authors.initials: RS
authors.lastname: Birolo
authors.forename: Nicole
authors.initials: N
authors.lastname: Carballido-Perrig
authors.forename: Francesca
authors.initials: F
authors.lastname: Ficara
authors.forename: Cristina
authors.initials: C
authors.lastname: Mocchetti
authors.forename: Jonathan
authors.initials: J
authors.lastname: Dando
authors.forename: José M
authors.initials: JM
authors.lastname: Carballido
authors.forename: Claudio
authors.initials: C
authors.lastname: Bordignon
authors.forename: Maria Grazia
authors.initials: MG
authors.lastname: Roncarolo
authors.forename: Marco
authors.initials: M
authors.lastname: Bregni
authors.forename: Alessandro
authors.initials: A
authors.lastname: Aiuti
mesh_terms: D000818:Animals; D018952:Antigens, CD34 / Q000032:analysis* / Q000378:metabolism; D003114:Colony-Forming Units Assay; D005434:Flow Cytometry; D005822:Genetic Vectors; D019650:Hematopoietic Stem Cell Mobilization; D006412:Hematopoietic Stem Cells / Q000737:chemistry / Q000502:physiology*; D008214:Lymphocytes / Q000378:metabolism; D038041:Lymphopoiesis; D051379:Mice; D016513:Mice, SCID; D022423:Myeloid Cells / Q000378:metabolism; D020800:Receptor, Nerve Growth Factor / Q000235:genetics* / Q000276:immunology; D012190:Retroviridae / Q000235:genetics; D014161:Transduction, Genetic / Q000379:methods*
chemical_list: D018952:Antigens, CD34; D020800:Receptor, Nerve Growth Factor