I. Response to the Recommendations of the Subcommittee 
The Subcommittee made five recommendations for additional data; these fall 
into two categories: A) murine TIL animal model data; B) human TIL data. 
A) Murine TIL Animal Model 
1) Recommendations from the Subcommittee: 
a) Transfer of the vector to TIL from a suitable murine tumor system 
b) Detection of vector-marked TIL in recipient mice 
c) Analysis of retroviral replication, tumorigenesis and other 
undesired effects in recipient mice 
2) Introduction 
The Subcommittee suggested that a stipulation for approval of any human gene 
therapy protocol should be that the investigators acquire data-using the closest 
available animal model relevant to the human study being proposed. We agree with 
that position, and provide data using the soluble antigen- activated CD4 murine T 
lymphocyte system which is, we argue, the closest available animal model. 
Unfortunately, the animal model requested by the Subcommittee, the tumor antigen- 
activated murine T lymphocyte system (the murine TIL model) which are CD8 T 
cells, is not "available". As discussed below, these murine T lymphocytes are 
not transduced by our retroviral vectors. We have, therefore, carried out all 
the studies requested using soluble antigen- activated CD4 murine T lymphocytes 
which can be readily transduced with our vectors. Human tumor antigen- activated 
T lymphocytes (TIL) from some patients appear to be all CD4, from other patients 
all CD8 , while in other cases they are a mixture of both CD4 and CD8 . Both types 
of human T cells are readily transducable by the N2 vector. Therefore, neither 
the soluble antigen- activated CD4 murine system nor the tumor antigen- activated 
CD8 (TIL) murine system is exactly analogous to the human situation, and, for the 
purposes of our study, neither is superior to the other. Since we have obtained 
data using the closest available animal model, we feel that we have satisfied the 
spirit of the Subcommittee's recommendation. 
3) Data from the murine T lvmohocvte model svstem 
a) Murine soluble antigen- activated CD4 T lymphocytes are readily 
transduced with N2 based vectors just as are human tumor antigen- activated CD4 
and CD8 T lymphocytes. The critical question is l.b. above: can it be 
demonstrated in an animal model that vector-marked T lymphocytes can be detected 
in an animal after days or weeks of growth in vivo ? The answer is yes. In these 
experiments, the N2 vector was modified to contain the gene for human adenosine 
deaminase (ADA) in addition to the NeoR gene (the SAX vector) or the gene for rat 
growth hormone (PB2N vector). A B10.D2 murine CD4 T cell line specifically 
responsive to sperm whale myoglobin (line 14.1) was transduced with either vector 
[ 208 ] 
Recombinant DNA Research, Volume 13 
