C. HESDORFFER 4/93 
correct recombinational event in sufficient quantity. This is 
the major reason we favor the use of retroviruses to add a normal 
MDR gene with a strong promoter as the current method of choice 
to pursue the goal of gene therapy. Adenovirus and adeno- 
associated vectors (AAV) have also been used recently to transfer 
genes into cells (83,84). 
4 . RELEVANT PRELIMINARY STUDIES FROM OUR LABORATORY ; 
A. Transfer and Expression of Foreign Genes in Mice : 
We have developed unique retroviral ecotropic and 
amphotropic packaging lines for safe and optimal gene transfer 
(16,17). These packaging lines contain the gaq - pol and env genes 
on different plasmids to avoid recombination. The packaging cell 
lines that produce the highest titer ecotropic and amphotropic 
viruses are GP+E-86 and GP+ env AM-12 . respectively (16,17). These 
lines are both safe and highly efficient in gene transfer, and 
have been sent to >300 laboratories worldwide with no reports of 
wild-type virus production. Recently, GP+envAM-12 was approved 
by the RAC and the FDA for use in cancer patients with an IL-2 
gene-containing retroviral vector (85) . 
We have used the GP+E-86 packaging line to transfer the neo R gene 
in a retroviral vector (N2) into irradiated mice (24). In these 
studies, we demonstrated high-level gene transfer; -50% of the 
mice show neo R gene integration in spleen colonies 13 days 
posttransplantation by Southern blotting (24) . These spleen 
colonies are presumed to represent proliferation of individual 
early bone marrow cells in the mouse and may be a reflection of 
true multipotent bone marrow stem cells (86) . Transplantation of 
spleen colonies from neo R -transduced mice into secondary- 
irradiated recipients supports this conclusion (24) . Mouse bone 
marrow at later times also shows neo R gene integration. The 
number of sites of integration detected at late times is 1-5 
(24), consistent with previous reports suggesting that only a few 
stem cells are involved in repopulation (21,23,24,87,88). In 
addition, most or all of the hematopoietic cells in both spleen 
colonies and bone marrow contain the neo R gene as assessed by the 
intensity of the neo R signal on Southern blots as compared to a 
single copy (globin) gene (24) . Thus, a foreign gene can be 
stably transduced into the majority of bone marrow progeny in 
live mice. 
B. The MDR Gene as a Selectable Marker : 
We have shown that the human MDR gene can be 
successfully and stably transduced and expressed in mouse 
erythroleukemia cells (MELC) (47) and mouse bone marrow cells 
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