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Journal of Virology, September 2005, p. 10999-11013, Vol. 79, No. 17
0022-538X/05/$08.00+0     doi:10.1128/JVI.79.17.10999-11013.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

A Capsid-Modified Helper-Dependent Adenovirus Vector Containing the ß-Globin Locus Control Region Displays a Nonrandom Integration Pattern and Allows Stable, Erythroid-Specific Gene Expression

Hongjie Wang,¹ Dmitry M. Shayakhmetov,¹ Tobias Leege,^1, Michael Harkey,⁴ Qiliang Li,¹ Thalia Papayannopoulou,² George Stamatoyannopolous,¹ and André Lieber^1,3*

Division of Medical Genetics,¹ Division of Hematology, Department of Medicine,² Department of Pathology, University of Washington,³ Fred Hutchinson Cancer Research Center, Seattle, Washington 98195⁴

Received 18 April 2005/ Accepted 1 June 2005

Gene therapy for hemoglobinopathies requires efficient gene transfer into hematopoietic stem cells and high-level erythroid-specific gene expression. Toward this goal, we constructed a helper-dependent adenovirus vector carrying the ß-globin locus control region (LCR) to drive green fluorescent protein (GFP) expression, whereby the LCR-GFP cassette is flanked by adeno-associated virus (AAV) inverted terminal repeats (Ad.LCR-ß-GFP). This vector possesses the adenovirus type 35 fiber knob that allows efficient infection of hematopoietic cells. Transduction and vector integration studies were performed in MO7e cells, a growth factor-dependent CD34⁺ erythroleukemic cell line, and in cord blood-derived human CD34⁺ cells. Stable transduction of MO7e cells with Ad.LCR-ß-GFP was more efficient and less subject to position effects and silencing than transduction with a vector that did not contain the ß-globin LCR. Analysis of integration sites indicated that Ad.LCR-ß-GFP integration in MO7e cells was not random but tethered to chromosome 11, specifically to the globin LCR. More than 10% of analyzed integration sites were within the chromosomal ß-globin LCR. None of the Ad.LCR-ß-GFP integrations occurred in exons. The integration pattern of a helper-dependent vector that contained X-chromosomal stuffer DNA was different from that of the ß-globin LCR-containing vector. Infection of primary CD34⁺ cells with Ad.LCR-ß-GFP did not affect the clonogenic capacity of CD34⁺ cells. Transduction of CD34⁺ cells with Ad.LCR-ß-GFP resulted in vector integration and erythroid lineage-specific GFP expression.

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* Corresponding author. Mailing address: Division of Medical Genetics, University of Washington, Box 357720, Seattle, WA 98195. Phone: (206) 221-3973. Fax: (206) 685-8675. E-mail: lieber00{at}u.washington.edu.

Present address: Universität Greifswald, D-17487 Greifswald, Germany.

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Journal of Virology, September 2005, p. 10999-11013, Vol. 79, No. 17
0022-538X/05/$08.00+0     doi:10.1128/JVI.79.17.10999-11013.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

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