title: High-efficiency transduction of human lymphoid progenitor cells and expression in differentiated T cells.
abstract: Gene therapy strategies for humans have been limited by low transduction efficiencies and poor expression of retroviral vectors in differentiated progeny cells carrying the transduced vector. Here we describe a strategy utilizing a cell surface reporter gene, murine thy-1.2, selectable by fluorescence-activated cell sorting (FACS), to achieve higher gene marking efficiencies. Human CD34-positive cells were transduced by a murine retroviral vector bearing the thy-1.2 marker and pseudotyped with vesicular stomatitis virus G protein, followed by FACS to enrich for CD34-positive cells that express Thy-1.2 on the cell surface. Gene marking and expression after differentiation into thymocytes were assessed in a SCID-hu Thy/Liv mouse model for human lymphoid progenitor cell gene therapy. We found that virtually all of the differentiated T-cell progeny were marked with vector sequences. It is of particular importance that reconstitution with the selected cells resulted in expression of Thy-1.2 in up to 71% of donor-derived thymocytes. It is of note that the donor-derived thymocytes that did not express Thy-1.2 still harbored vector thy-1.2 sequences, suggesting repression of transgene expression in some cells during progenitor cell differentiation into thymocytes. These studies provide a proof of concept for efficient expression of transgenes through T-lymphoid differentiation and a potential basis for utilizing similar strategies in human gene therapy clinical trials.
authors.affiliation: Department of Microbiology & Immunology, UCLA School of Medicine, Los Angeles, California 90095-1678, USA.
authors.forename: D S
authors.initials: DS
authors.lastname: An
authors.forename: Y
authors.initials: Y
authors.lastname: Koyanagi
authors.forename: J Q
authors.initials: JQ
authors.lastname: Zhao
authors.forename: R
authors.initials: R
authors.lastname: Akkina
authors.forename: G
authors.initials: G
authors.lastname: Bristol
authors.forename: N
authors.initials: N
authors.lastname: Yamamoto
authors.forename: J A
authors.initials: JA
authors.lastname: Zack
authors.forename: I S
authors.initials: IS
authors.lastname: Chen
mesh_terms: D000818:Animals; D018952:Antigens, CD34; D005434:Flow Cytometry; D015870:Gene Expression; D015316:Genetic Therapy; D005822:Genetic Vectors; D006412:Hematopoietic Stem Cells; D006801:Humans; D051379:Mice; D013601:T-Lymphocytes; D018800:Thy-1 Antigens / Q000235:genetics*
chemical_list: D018952:Antigens, CD34; D018800:Thy-1 Antigens