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High-frequency cell surface expression of a foreign protein in murine
hematopoietic stem cells using a new retroviral vector
DB Tumas, GJ Spangrude, DM Brooks, CD Williams and B Chesebro
Laboratories of Persistent Viral Diseases, National Institute of Allergy
and Infectious Diseases, National Institutes of Health, Rocky Mountain
Laboratories, MT 59840, USA.
A retroviral vector (pSFF) derived from murine Friend spleen focus forming
virus was used to transduce murine hematopoietic stem cells and express a
cell surface marker protein, mutated murine prion protein, in vitro and in
vivo after transplantation. To enhance retroviral vector integration in
bone marrow cells, mice were treated with 5-fluorouracil (5-FU) to increase
stem cell mitotic activity, which peaked on day 8 post-5-FU. The
infectivity titer of the vector, pSFF-mPrP-3F4, was determined by a novel
assay in which antigen-positive foci of infected cells were detected after
replication and spread of the vector in cultures of mixed packaging cell
lines. Infection of Sca-1+/Lineageneg- low cells with pSFF-mPrP-3F4
resulted in marker protein expression in 40% of the progeny cells after 7
days of culture. Transplantation of marrow cells or sorted
Sca-1+/Lineageneg-low cells transduced with vector resulted in 3F4-positive
mPrP expression in 11% to 37% of donor- derived peripheral blood leukocytes
at 2 weeks. Though the percentage of 3F4-positive blood cells gradually
declined, at 28 weeks 23% of recipient mice still maintained expression of
the marker gene. Expression was observed in lymphoid, myeloid, and
erythroid lineages and was detected in Sca-1+/Lineageneg-low marrow cells.
The multilineage, high-frequency expression observed suggests that pSFF may
be useful in gene therapy directed at hematopoietic stem cells and their
differentiated progeny.
Volume 87,
Issue 2,
pp. 509-517,
01/15/1996
Copyright © 1996 by The American Society of Hematology
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