Using a murine bone marrow transplantation model, we evaluated the lon
g-term engraftment of retrovirally transduced bone marrow cells in non
myeloablated hosts. Male bone marrow was stimulated in a cocktail of i
nterleukin-3 (IL-3), IL-6, IL-11, and stem cell factor (SCF) for 48 ho
urs, then cocultured on the retroviral producer line MDR18.1 for an ad
ditional 24 hours. Functional transduction of hematopoietic progenitor
s was detected in vitro by reverse transcriptase-polymerase chain reac
tion (RT-PCR) amplification of multiple drug resistance 1 (MDR1) mRNA
from high proliferative potential-colony forming cell (HPP-CFC) coloni
es. After retroviral transduction, male bone marrow cells were injecte
d into nonablated female mice. Transplant recipients received three TA
XOL (Bristol-Myers, Princeton, NJ) injections (10 mg/kg) over a 14-mon
th period. Transplant recipient tissues were analyzed by Southern blot
and fluorescence in situ hybridization for Y-chromosome-specific sequ
ences and showed donor cell engraftment of approximately 9%. However,
polymerase chain reaction amplification of DNAs from bone marrow, sple
en, and peripheral blood showed no evidence of the transduced MDR1 gen
e. RT-PCR analysis of total bone marrow RNA showed that transcripts fr
om the MDR1 gene were present in a fraction of the engrafted donor cel
ls. These data show functional transfer of the MDR1 gene into nonmyelo
ablated murine hosts. However, the high rates of in vitro transduction
into HPP-CFC, coupled with the low in vivo engraftment rate of donor
cells containing the MDR1 gene, suggest that the majority of stem cell
s that incorporated the retroviral construct did not stably engraft in
the host. Based on additional studies that indicate that ex vivo cult
ure of bone marrow induces an engraftment defect concomitantly with pr
ogression of cells through S phase, we propose that the cell cycle tra
nsit required for proviral integration reduces or impairs the ability
of transduced cells to stably engraft. (C) 1997 by The American Societ
y of Hematology.