Adenoviral vector design for high-level transgene expression in primitive human hematopoietic progenitors
Open Access
- 1 December 2000
- journal article
- research article
- Published by Springer Nature in Gene Therapy
- Vol. 7 (24) , 2132-2138
- https://doi.org/10.1038/sj.gt.3301352
Abstract
Adenoviral vector-mediated transient gene expression can provide new possibilities for ex vivo manipulation of quiescent hematopoietic stem cells (HSC). In order to define a suitable expression cassette for high levels of transgene expression in HSCs, we have studied the level of transgene expression in human CD34+CD38− cells using adenoviral vectors with various gene expression cassettes encoding the enhanced green fluorescence protein (EGFP) gene. CD34+ hematopoietic cells were cultured in serum-free medium with megakaryocyte growth and development factor (MGDF) alone for supporting the survival of primitive progenitors or with MGDF, c-kit ligand (KL) and flt3 ligand (FL) for inducing proliferation of primitive progenitors. With all the vectors tested, higher percentages of EGFP expressing cells were found in CD34+CD38− cells than those in CD34+CD38high cells from all donors tested. The phosphoglycerate kinase (PGK)-1 promoter was found to allow higher levels of EGFP expression than the human cytomegalovirus (HCMV) promoter in CD34+CD38− cells. Replacing the SV40 polyadenylation signal with the human β-globin gene IVS2 and polyadenylation signal in the expression cassette (Ad5xPGK-EGFP-β-globin) enhanced the level of EGFP expression markedly further. These results provide a guideline for the development of adenoviral vectors for gene expression in human primitive hematopoietic progenitor cells.Keywords
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