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Prepublished online as a Blood First Edition Paper on October 17, 2002; DOI 10.1182/blood-2002-07-2238. This Article Full Text (PDF) All Versions of this Article: 2002-07-2238v1 101/4/1284    most recent Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Rights and Permissions Citing Articles Citing Articles via HighWire Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Woods, N.-B. Articles by Karlsson, S. Search for Related Content PubMed PubMed Citation Articles by Woods, N.-B. Articles by Karlsson, S. Social Bookmarking                   What's this? Submitted July 25, 2002 Accepted September 6, 2002 Lentiviral vector transduction of NOD/SCID repopulating cells results in multiple vector integrations per transduced cell: risk of insertional mutagenesis Niels-Bjarne Woods, Arne Muessig, Manfred Schmidt, Johan Flygare, Karin Olsson, Patrick Salmon, Didier Trono, Christof von Kalle*, and Stefan Karlsson Department of Molecular Medicine and Gene Therapy, Lund University, Lund, Sweden Dept. I of Internal Medicine, University of Freiburg, Freiburg, Germany Institute for Molecular Medicine and Cell Research, University of Freiburg, Freiburg, Germany Department of Genetics and Microbiology, University of Geneva Medical School, Geneva, Switzerland Program for Molecular and Gene Therapy, Cincinnati Children's Research Foundation, Cincinnati, OH, USA * Corresponding author; email: Christof.Kalle{at}chmcc.org. Efficient vector transduction of hematopoietic stem cells is a requirement for successful gene therapy of hematological disorders. We asked whether human umbilical cord blood CD34+ CD38lo NOD/SCID repopulating cells (SRC) could be efficiently transduced using lentiviral vectors, with a particular focus on the average number of vector copies integrating into the these primitive progenitor cells. Mouse bone marrow was analyzed by FACS and by semi-quantitative PCR to determine the transduction efficiency into SRC. Lentiviral vector transduction resulted in an average of 22% (range 3-90%) of the human cells expressing GFP, however, multiple vector copies were present in human hematopoietic cells, with an average of 5.6±3.3 (n=12) copies per transduced cell. To confirm the ability of lentiviral vectors to integrate multiple vector copies into SRC, linear amplification mediated (LAM)-PCR was used to analyze the integration site profile of a selected mouse showing low level engraftment and virtually all human cells expressing GFP. Individually picked CFU-GM colonies derived from the bone marrow of this mouse were analyzed and shown to have the same 5 vector integrants within each colony. Interestingly, one integration site of the five that were sequenced in this mouse was located in a known tumor-suppressor gene, BRCA1. Therefore, these findings demonstrate the ability of lentiviral vectors to transduce multiple copies into a subset of NOD/SCID repopulating cells. While this is efficient in terms of transduction and transgene expression, it may increase the risk of insertional mutagenesis. CiteULike    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this? This article has been cited by other articles: N. Ali, C. Karlsson, M. Aspling, G. Hu, N. Hacohen, D. T. Scadden, and J. Larsson Forward RNAi screens in primary human hematopoietic stem/progenitor cells Blood, April 16, 2009; 113(16): 3690 - 3695. [Abstract] [Full Text] [PDF] E. Kieback, J. Charo, D. Sommermeyer, T. Blankenstein, and W. 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	Copyright © 2002 by American Society of Hematology         Online ISSN: 1528-0020