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TISSUE-SPECIFIC STEM CELLS

Sustained Long-Term Engraftment and Transgene Expression of Peripheral Blood CD34⁺ Cells Transduced with Third-Generation Lentiviral Vectors

^aLaboratory of Clinical Oncology, Department of Oncological Sciences, University of Torino Medical School, Institute for Cancer Research and Treatment, Candiolo (Torino) Italy;
^cDepartment of Public Health and Microbiology, University of Torino Medical School, Torino, Italy;
^bPediatric Onco-Haematology Unit, Stem Cell Transplantation and Cellular Therapy Center, Regina Margherita Children's Hospital, Torino, Italy;
^dHematology and Clinical Immunology Section and
^eInternal Medicine and Oncological Science Section, University of Perugia, Perugia, Italy

Key Words. Ex vivo gene transfer • Hematopoietic stem cells • Enhanced green fluorescent protein • Expansion • Mobilized peripheral blood

Correspondence: Wanda Piacibello, M.D., Laboratory of Clinical Oncology, IRCC Institute for Cancer Research and Treatment, Provinciale 142, 10060 Candiolo, Torino, Italy. Telephone: 390119933349; Fax: 390119933522; e-mail: wanda.piacibello{at}ircc.it

Received February 19, 2008; accepted for publication March 14, 2008.
First published online in STEM CELLS EXPRESS   March 27, 2008.

As mobilized peripheral blood (MPB) represents an attractive cell source for gene therapy, we investigated the ability of third-generation lentiviral vectors (LVs) to transfer the enhanced green fluorescent protein gene into MPB CD34⁺ cells in culture conditions allowing expansion of transplantable human hematopoietic stem cells. To date, few studies have reported transduction of MPB cells with vesicular stomatitis virus G pseudotyped LVs. The critical issue remains whether primitive, hematopoietic repopulating cells have, indeed, been transduced. In vitro (5 weeks' culture in FLT3 ligand + thrombopoietin + stem cell factor + interleukin 6) and in vivo (serial transplantation in NOD/SCID mice) experiments show that MPB CD34⁺ cells can be effectively long-term transduced by LV and maintain their proliferation, self-renewal, and multilineage differentiation potentials. We show that expansion following transduction improves the engraftment of transduced MPB CD34⁺ (4.6-fold expansion of SCID repopulating cells by limiting dilution studies). We propose ex vivo expansion after transduction as an effective tool to improve gene therapy protocols with MPB.

Disclosure of potential conflicts of interest is found at the end of this article.