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Retroviral transduction of quiescent haematopoietic cells using a packaging cell line expressing the membrane-bound form of stem cell factor

Gene Therapy volume 6, pages 1084–1091 (1999)Cite this article

Abstract

Gene therapy vectors based on murine retroviruses are unable to transduce non-dividing cells. This has proven a particular problem in the haematopoietic system where the target cells of choice, the pluripotent stem cells are quiescent. In an attempt to circumvent this difficulty we have constructed a retroviral producer line that expresses the membrane bound form of human recombinant stem cell factor (SCF) on its cell surface. This should enable the retroviral producers to deliver a growth signal to the target cells simultaneous with their exposure to retrovirus. We tested the ability of these modified producers to transduce a growth factor-starved, SCF-dependent cell line (TF-1) and demon- strated that these cells, though quiescent, can still be successfully transduced. This approach was extended to targeting of umbilical cord blood CD34+ cells, a predominantly quiescent population that normally require the addition of cytokines for efficient transduction. Using the SCF-expressing producer line in the absence of exogenously added cytokines, we observed a marked stimulation in transduction efficiency over that achieved using the parent producer line alone. Colonies derived from these cells arising in semi-solid media were also shown to be positive for expression of a retrovirally encoded reporter gene.

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Acknowledgements

We would like to thank Dr Colin Porter (ICR, London) for providing the NIH3T3 and AM12(lacZ) cell lines and Joanna Povey for the gift of plasmid pREP8-ΔSCF. Thanks are also due to Professor David Linch (UCLMS, London) and Immunex, Seattle, WA, USA for their kind gifts of GM-CSF and SCF cDNA, respectively. This work was supported by grants from the MRC and the CGD Research Trust.

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  1. Department of Haematology, Imperial College School of Medicine, St Mary’s Campus, Norfolk Place, London, W2 1PG, UK

    A Sehgal, N Weeratunge & C Casimir

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  1. A Sehgal
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  2. N Weeratunge
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Sehgal, A., Weeratunge, N. & Casimir, C. Retroviral transduction of quiescent haematopoietic cells using a packaging cell line expressing the membrane-bound form of stem cell factor. Gene Ther 6, 1084–1091 (1999). https://doi.org/10.1038/sj.gt.3300932

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