lasting from one that will be short-lived? Could such a characteristic be 
exploited to increase TIL survival in vivo ? How long does an infused TIL 
population remain in the body? At what sites? Are there TIL that survive but 
nonetheless lose their effectiveness? Are there characteristics that could 
predict this behavior? 
In order to obtain answers to these questions it would be necessary to have 
a means to efficiently mark TIL in a manner such that they could be identified 
and re- isolated from a patient and studied. At present, there are no reliable 
clinical procedures for long-term marking of autologous cells in vivo . 
Radionuclides have been extensively employed as labels to permit studies of a 
cell population's distribution and survival in the body but have several 
properties that limit their practical effectiveness. Rapid decay of the commonly 
utilized isotopes prevents their use for long-term studies and the dose of 
radiation exposure received if longer-lived isotopes are employed is unacceptable 
for clinical studies. In addition, the loss of label or its reutilization or 
sequestration at sites no longer related to the original "marked" call is a 
significant problem for some types of long-term cell survival studies. 
The use of the radionuclide Indium^^^ for short-term labeling of TIL has 
been useful and results using this technique are described below in the Proeress 
Report (Section III. A). 
We propose to use a totally new procedure for marking TIL for long-term 
studies, namely, to insert a selectable marker gene via retroviral-mediated gene 
transfer. As a permanent and stable component of the TIL genome, the TIL and all 
its offspring would be marked in such a way that these calls could always be 
identified and re- isolated even if they represented only a minute fraction of the 
total cells present (theoretically, even one cell in a biopsy specimen could be 
isolated, expanded, and studied). The laboratories of Dr. Anderson and Dr. 
Blaese have extensive experience with retroviral-mediated gene transfer into many 
types of cells including lymphocytes (1-3) and bone marrow cells (4). They also 
have considerable experience with In vivo gene transfer and expression in a 
number of species ranging from mice (5) to primates (6) . 
We have already demonstrated that TIL from several different patients can be 
easily transduced via retroviral-mediated gene transfer and that the transferred 
selectable marker gene is efficiently expressed (see details in the Progress 
Report . Section III.B.). 
The use of retroviral-mediated gene transfer as a means for marking TIL in 
the present Project has several practical advantages as well as a number of 
theoretical risks. Retroviral-mediated gene transfer has not previously been used 
in man and therefore the risks and benefits of this proposed technology need to 
be analyzed carefully.. The recombinant DNA aspects of this Addition, therefore, 
will be reviewed by the Recombinant DNA Advisory Committee (RAC) as well as by 
the Human Gene Therapy Subcommittee of the RAC in open public hearings. Their 
recommendation is then referred to the Director of NIH for final approval. The 
FDA must also approve the use of a biological agent (the recombinant retroviral 
vector). No studies will be carried out on patients until approval is received 
from all these quarters. 
Recombinant DNA Research, Volume 14 
[3] 
