Human Gene Therapy Subcommittee - 11/21-22/91 
G418 or PCR analysis to determine repopulation with neo R transduced leukemic cells. 
Dr. Cornetta stated that both of these assays will be used. He explained that colonies 
would be picked and assayed over time for the marker gene because it is unknown 
whether the LNL6 vector will be expressed long term. Non-G418 selected colonies will 
be assayed for the presence of the marker gene. 
Dr. Leventhal was concerned that patients will be requested to have multiple bone 
marrow collections because the investigators will not know when the patient has 
relapsed. The protocol has to been designed to collect and examine bone marrow at 
optimum time points. Dr. Cornetta responded that he will include a revised time 
schedule for bone marrow collection if requested to do so by the RAC. Dr. B. Murray 
asked if the time of relapse could be anticipated in most of the proposed patient 
population. Dr. Cornetta said that the time of relapse could be estimated, and the 
protocol will be revised to reflect bone marrow collection just prior to relapse. 
Dr. Leventhal inquired how marked leukemic cells will be distinguished from marked 
normal cells since the frequency of marking will be approximately equal in both 
populations of cells. Dr. Cornetta explained that it is critical to grow cells in 
methylcellulose culture in order to distinguish between normal and leukemic cells. PCR 
will be performed on the GM colonies to determine transduction efficiency. The GM 
colonies will then be sorted by FACS into normal and blast populations. 
Dr. Parkman asked if there is an exclusion criterion for the degree of transduction that 
will be required in the bone marrow prior to transplantation. Dr. Cornetta responded 
that there is no exclusion criterion. Dr. Parkman was concerned that if the number of 
transduced cells was too low, the likelihood of obtaining an informative outcome would 
decrease. Dr. Cornetta agreed that a minimum level of bone marrow cell marking would 
be an acceptable criterion to include in the protocol. 
Dr. Cornetta introduced his co-investigator, Dr. Tricot, to respond to questions regarding 
the transplantation process. Dr. Leventhal asked about the one-year relapse incidence in 
ALL and AML patients who have received autologous bone marrow transplantation and 
who are in second remission. Dr. Tricot answered that the relapse rate is approximately 
50% in AML patients; no ALL patients have received autologous transplants. Dr. 
Leventhal asked if 50% of the patients who have received transplants relapsed or if 50% 
of the patients who lived to a year did not relapse. Dr. Tricot stated that 50% referred 
to the number of patients who survived one year with no relapse. Dr. Leventhal said 
that the investigators need to know how many patients will be available for this study 
since a proportion of these transplant patients invariably die from sepsis or graft-versus- 
host (GVH) disease. Dr. Tricot stated that the majority of autologous transplant patients 
fail exclusively due to relapse, and that other complications are rare. Therefore, 
approximately 5 patients out of a starting population of 10 patients would be eligible for 
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