The presence of 12,000 marked leukemic marrow blasts and 
60,000 peripheral blood leukemic blasts in the autologous infusion 
is clearly sufficient to generate a polyclonal relapse, if relapse 
occurs from cells in the autologous cells infused. The presence of 
such marked blasts would be detectable by PCR if relapse occurred 
from leukemic blast cells present in the autologous infusion, as 
the ratio of marked leukemic blasts/non-blasts cells would clearly 
be higher at relapse than 1/300,000. 
These data show that the probability of detecting a polyclonal 
relapse with marked cellsk, if it arises from residual leukemic 
blastic cells in the autologous infusion, is very high. 
The animal model data developed by Drs. Rosenberg and Anderson 
for the LNL6 vector was outlined in the original N2-TIL cell 
protocol previously approved by the NIH RAC committee. The safety 
of the LNL6 vector was established in those studies. Furthermore, 
these studies have shown that the introduction of the LNL6 vector 
does not change the phenotype of the TIL cells. Those studies have 
shown that there is no horizontal infectious spread following 
infection of the TIL cells by LNL6 (1) . The ways in which the 
animal models were similar to the human system were summarized in 
the previous LNL6 NEO TIL protocol. 
The cell culture methods used to assess the in vitro efficacy 
of the gene transfer system are described above. The conditions 
are identical to those proposed for the treatment of human marrow 
in the clinical setting. 
The use of the direct methylcellulose method is similar to the 
human system in that it can measure the efficiency of transfer and 
expression (methylcellulose) of the LNL6 retroviral genome. The 
differences include: 
The number of cycling cells in the methylcellulose and in vivo 
settings, and the length of time available for the retention of 
the LNL6 retroviral genome may be different. 
The difference between the present protocol and the LNL6 TIL 
cell experiments is that the TIL cell experiments are short-term 
experiments, and do not require engraftment. In addition, a 
mixture of leukemia and diploid cells are used in our experiments. 
However, our protocol for CML is similar to one approved for AML 
(Malcolm Brenner submission, 11/90) , which also uses human 
autologous cells which are comprised of a mixture of normal and 
leukemic, rather than purified normal lymphocytes as in the TIL 
cell protocol. 
A previous protocol (by Dr. Malcolm Brenner) which has been 
approved by the RAC on 2/4/91 addressed the stability of the 
transgenome. 
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Recombinant DNA Research, Volume 14 
