CNS 18 - 7 
cells maintain some mitotic activity, and may be at some (minimal) risk for viral 
incorporation. Therefore, the likelihood of specific transduction of desired gene 
into tumor cells vs other cells is enhanced. Second, the brain is a partially 
immunologically privileged site which should allow a somewhat longer survival 
of the xenogeneic murine cells in the brain, leading to greater virus production 
in situ and thus potentially a greater transduction efficiency than might be seen 
in other organs. This and the ability of many CNS tumors to depress local 
immunity (12) should allow for the transduction of greater numbers of tumor 
cells. By design, this period of survival will be limited as cells that integrate and 
express the HS-tkgene, including the retroviral vector producer cells, should later 
be destroyed by GCV. 
2.3.1 Gene Transfer Methods 
There are 3 major methods of gene transfer: 1) chemical, 2) 
physical and 3) viral (e.g. herpes simplex, adenovirus, retrovirus) 
(13). Physical and chemical methods have poor gene integration 
efficiency, ranging from 1:1,000, to 1:100,000, making them 
impractical for most clinical applications. The adenovirus and 
herpes simplex viruses have the ability to replicate extra- 
chromosomally, and have been used as vectors to transfer genes 
into cells where they replicate in an extrachromosomal location. 
Therefore, these viruses, unlike the murine retrovirus from which 
our vector is derived, can transfer genes into non-replicating 
tissues (e.g. lung, brain). 
2.3.2 Retroviral-Mediated Gene Transfer (14) 
Murine retroviral vectors are proven extremely efficient for gene 
transfer into mammalian cells - as high as 90% in cultured murine 
fibroblast line(s). Murine retroviral vectors integrate and 
subsequently express their genes at high efficiency only in 
proliferating cells. Because tumor cells are the predominant 
mitotic cell in the otherwise post-mitotic CNS, differential 
transduction of tumor should be maximal while transduction into 
brain parenchymal cells is minimal or absent. The moloney 
murine leukemia virus-based (MoMLV) vectors to be used in the 
current protocol have been designed to minimize the possibility of 
recombination leading to regeneration of replication-competent 
virus (15,16). 
2.3.3 Experience with Retroviral- mediated Gene Transfer Into Humans 
N2/TIL Marking Study: 
In 1989, the first gene transfer experiment in humans was 
conducted at the NIH. This study involved the treatment of 10 
patients with autologous T-cells (TIL) transduced with the 
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