IMPROVED RETROVIRAL GENE-TRANSFER INTO MURINE AND RHESUS PERIPHERAL-BLOOD OR BONE-MARROW REPOPULATING CELLS PRIMED IN-VIVO WITH STEM-CELL FACTOR AND GRANULOCYTE-COLONY-STIMULATING FACTOR
Ce. Dunbar et al., IMPROVED RETROVIRAL GENE-TRANSFER INTO MURINE AND RHESUS PERIPHERAL-BLOOD OR BONE-MARROW REPOPULATING CELLS PRIMED IN-VIVO WITH STEM-CELL FACTOR AND GRANULOCYTE-COLONY-STIMULATING FACTOR, Proceedings of the National Academy of Sciences of the United Statesof America, 93(21), 1996, pp. 11871-11876
In previous studies we showed that 5 days of treatment with granulocyt
e colony-stimulating factor (G-CSF) and stem cell factor (SCF) mobiliz
ed murine repopulating cells into the peripheral blood (PB) and that t
hese cells could be efficiently transduced with retroviral vectors. We
also found that, 7-14 days after cytokine treatment, the repopulating
ability of murine bone marrow (BM) increased 10-fold. In this study w
e examined the efficiency of gene transfer into cytokine-primed murine
BR4 cells and extended our observations to a nonhuman primate autolog
ous transplantation model. G-CSF/SCF-primed murine BM cells collected
7-14 days after cytokine treatment were equivalent to post-5-fluoroura
cil BM or G-CSF/SCF-mobilized PB cells as targets for retroviral gene
transfer. In nonhuman primates, CD34-enriched PB cells collected after
5 days of G-CSF/SCF treatment and CD34-enriched BM cells collected 14
days later were superior targets for retroviral gene transfer. When a
clinically approved supernatant infection protocol with low-titer vec
tor preparations was used, monkeys had up to 5% of circulating cells c
ontaining the vector for up to a year after transplantation. This rela
tively high level of gene transfer was confirmed by Southern blot anal
ysis. Engraftment after transplantation using primed BM cells was more
rapid than that using steady-state bone marrow, and the fraction of B
M cells having the most primitive CD34(+)/CD38(-) or CD34(+)/CD38(dim)
phenotype increased 3-fold. We conclude that cytokine priming with G-
CSF/SCF may allow collection of increased numbers of primitive cells f
rom both the PB and BM that have improved susceptibility to retroviral
transduction, with many potential applications in hematopoietic stem
cell-directed gene therapy.