culture (40,41), there is a well established CTL involvement (40,41), many melanomas have low 
Class 1 MHC levels (42-44), and they respond to IL-2/LAK/TIL and a-DFN therapies (10). The 
patient population will be melanoma patients with metastatic disease that has failed other forms of 
therapy. 
3.0 PRE-CLINICAL STUDIES 
3.1 SUMMARY 
The feasibility of the approach has been tested in several murine tumor model systems using 
retroviral vectors that express murine y-IFN. Gene-modified murine tumor cell lines secreted y- 
IFN and exhibited substantial increases in the surface expression of Class I MHC proteins. The 
modified tumor cells were substantially less tumorigenic, apparently due to their ability to invoke 
increased anti-tumor immunity. Further, injection of y-IFN-expressing tumor cells resulted in 
potent CTL activity capable of specifically lysing the unmodified parent tumor cell. Unmodified 
tumor cells did not engender such a response. Pre-injection with irradiated y-IFN-expressing 
tumor cells, but not the unmodified tumor cells, protected animals from subsequent challenge with 
the unmodified tumor. This effect is presumably due to the observed induction of the tumor 
specific CTL. These data show that the naturally poor immunogenic properties of tumor cells can 
be improved by the constitutive expression of y-IFN delivered by retroviral vectors. The immune 
system, thus primed, can mount potent responses against the unmodified parent tumor. This 
conclusion suggests that a similar gene transfer based immunotherapeutic approach to human 
cancer may be of substantial benefit to cancer patients. 
A high titer replication incompetent amphotropic retroviral vector producing cell line which can be 
used to deliver the human y-IFN gene to a variety of cell types has been generated. We have used 
this vector producing cell line to introduce the human y-IFN gene into a number of human 
melanoma cell lines. Transduction of human melanoma cells by amphotropic retroviral vectors is 
efficient and results in the rapid expression of biologically active human y-IFN. Transduced tumor 
cell lines show increased expression of human Class I and II MHC (HLA) as determined by 
protein immunoblotting, immunofluorescence, and fluoresence activated cell sorting (FACS) 
analysis. y-IFN-transduced melanoma cells can stimulate CTLs from melanoma patients 5-40X 
more efficiently than unmodified tumor cells. 
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Recombinant DNA Research, Volume 17 
