Abstract
One of the main impediments to effective gene therapy of blood disorders is the resistance of human hematopoietic stem cells to stable genetic modification. We show here that a small minority of retrovirally transduced stem cells can be selectively enriched in vivo, which might be a way to circumvent this obstacle. We constructed two retroviral vectors containing an antifolate-resistant dihydrofolate reductase cDNA transcriptionally linked to a reporter gene. Mice were transplanted with transduced bone marrow cells and then treated with an antifolate-based regimen that kills unmodified stem cells. Drug treatment significantly increased the percentage of vector-expressing peripheral blood erythrocytes, platelets, granulocytes, and T and B lymphocytes. Secondary transplant experiments demonstrated that selection occurred at the level of hematopoietic stem cells. This system for in vivo stem-cell selection provides a means to increase the number of genetically modified cells after transplant, and may circumvent an substantial obstacle to successful gene therapy for human blood diseases.
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Acknowledgements
The authors thank E. Allay for her technical assistance, E. Wingfield and A. M. Hamilton-Easton for their assistance with the flow cytometry analyses, and A. Nienhuis for critical review of the manuscript. We also thank AMGEN (Thousand Oaks, California) for their gift of rat SCF and human IL-6. This work was supported in part by National Heart, Lung, and Blood Institute Program Project Grant No. P01 HL 53749 (BPS), The James S. McDonnell Foundation Grant No. 94-50 (BPS), The John H. Sununu Postdoctoral Fellowship (JAA), The Assisi Foundation of Memphis Grant 94-00, US Public Health Service Grant No. P01 CA 31922 (RLB), Cancer Center Support Grant No. P30 CA 21765 (BPS, RLB), and the American Lebanese Syrian Associated Charities (ALSAC)
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Allay, J., Persons, D., Galipeau, J. et al. In vivo selection of retrovirally transduced hematopoietic stem cells. Nat Med 4, 1136–1143 (1998). https://doi.org/10.1038/2632
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DOI: https://doi.org/10.1038/2632
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