Transduction of pluripotent human hematopoietic stem cells demonstrated by clonal analysis after engraftment in immune-deficient mice.
- 19 March 1996
- journal article
- Published by Proceedings of the National Academy of Sciences in Proceedings of the National Academy of Sciences
- Vol. 93 (6) , 2414-2419
- https://doi.org/10.1073/pnas.93.6.2414
Abstract
Gene transduction of pluripotent human hematopoietic stem cells (HSCs) is necessary for successful gene therapy of genetic disorders involving hematolymphoid cells. Evidence for transduction of pluripotent HSCs can be deduced from the demonstration of a retroviral vector integrated into the same cellular chromosomal DNA site in myeloid and lymphoid cells descended from a common HSC precursor. CD34+ progenitors from human bone marrow and mobilized peripheral blood were transduced by retroviral vectors and used for long-term engraftment in immune-deficient (beige/nude/XIS) mice. Human lymphoid and myeloid populations were recovered from the marrow of the mice after 7-11 months, and individual human granulocyte-macrophage and T-cell clones were isolated and expanded ex vivo. Inverse PCR from the retroviral long terminal repeat into the flanking genomic DNA was performed on each sorted cell population. The recovered cellular DNA segments that flanked proviral integrants were sequenced to confirm identity. Three mice were found (of 24 informative mice) to contain human lymphoid and myeloid populations with identical proviral integration sites, confirming that pluripotent human HSCs had been transduced.Keywords
This publication has 19 references indexed in Scilit:
- Analysis of optimal conditions for retroviral-mediated transduction of primitive human hematopoietic cellsBlood, 1995
- Lack of expression from a retroviral vector after transduction of murine hematopoietic stem cells is associated with methylation in vivo.Proceedings of the National Academy of Sciences, 1994
- Gene marking to determine whether autologous marrow infusion restores long-term haemopoiesis in cancer patientsThe Lancet, 1993
- Treatment of Severe Combined Immunodeficiency Disease (SCID) due to Adenosine Deaminase Deficiency with CD34+Selected Autologous Peripheral Blood Cells Transduced with a Human ADA Gene (Amendment). National Institutes of HealthHuman Gene Therapy, 1993
- Long-term expression of human adenosine deaminase in rhesus monkeys transplanted with retrovirus-infected bone-marrow cells.Proceedings of the National Academy of Sciences, 1992
- Clonal proliferation of murine lymphohemopoietic progenitors in culture.Proceedings of the National Academy of Sciences, 1992
- Proliferation of totipotent hematopoietic stem cells in vitro with retention of long-term competitive in vivo reconstituting ability.Proceedings of the National Academy of Sciences, 1992
- Clonal and systemic analysis of long-term hematopoiesis in the mouse.Genes & Development, 1990
- IMPROVED RETROVIRAL VECTORS FOR GENE-TRANSFER AND EXPRESSION1989
- Introduction of a selectable gene into primitive stem cells capable of long-term reconstitution of the hemopoietic system of W/W miceCell, 1985