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Blood, 1 September 2007, Vol. 110, No. 5, pp. 1448-1457.
Prepublished online as a Blood First Edition Paper on April 24, 2007; DOI 10.1182/blood-2006-12-060814.
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Submitted December 5, 2006
Accepted April 20, 2007
Lentiviral vectors containing an enhancer-less
ubiquitously-acting chromatin opening element (UCOE)
provide highly reproducible and stable transgene
expression in haematopoietic cells
Fang Zhang, Susannah I Thornhill, Steven J Howe, Meera Ulaganathan, Axel Schambach, Joanna Sinclair, Christine Kinnon, H Bobby Gaspar, Michael Antoniou, and Adrian J. Thrasher*
Molecular Immunology Unit, Centre for Immunodeficiency, Institute of Child Health, University College London, London, United Kingdom
Department of Experimental Hematology, Hannover Medical School, Hannover, Germany
Department of Clinical Immunology, Great Ormond Street Hospital NHS Trust, London, United Kingdom
Department of Medical and Molecular Genetics, Nuclear Biology Group, Kings College London School of Medicine, London, United Kingdom
* Corresponding author; email: a.thrasher{at}ich.ucl.ac.uk.
Ubiquitously-acting chromatin opening elements (UCOEs) consist of methylation-free CpG islands encompassing dual, divergently transcribed promoters of housekeeping genes which have been shown to confer resistance to transcriptional silencing and to produce consistent and stable transgene expression in tissue culture systems. To develop improved strategies for haematopoietic cell gene therapy we have assessed the potential of the novel human HNRPA2B1-CBX3 UCOE (A2UCOE) within the context of a SIN lentiviral vector. Unlike viral promoters, the enhancer-less A2UCOE gave rise to populations of cells that expressed a reporter transgene at a highly reproducible level. The efficiency of expression per vector genome was also markedly increased in vivo compared to vectors incorporating either SFFV or CMV promoters, suggesting a relative resistance to silencing. Furthermore, an A2UCOE-IL2RG vector fully restored the IL-2 signalling pathway within IL2RG deficient human cells in vitro and successfully rescued the SCID-X1 phenotype in a mouse model of this disease. These data indicate that the A2UCOE displays highly reliable transcriptional activity within a lentiviral vector, largely overcoming insertion site position effects and giving rise to therapeutically relevant levels of gene expression. These properties are achieved in the absence of classical enhancer activity and therefore may confer a high safety profile.
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