Human Gene Therapy Subcommittee - 11/21-22/91 
this study due to disease relapse. 
Dr. Leventhal asked if the investigators will purge any of the patient's bone marrow for 
this protocol? Dr. Tricot replied that none of the bone marrow will be purged because 
purging increases the duration of aplasia by 15-30 days, and there is no convincing 
evidence that purging is superior to not purging the marrow. Dr. Leventhal asked if the 
conditioning regimen will be the same for all of the patients. Dr. Tricot replied that the 
same conditioning regimen will be employed for all AML and ALL patients. 
Dr. Gellert said that even when patients are in remission, there are large numbers of 
surviving leukemic cells. He inquired if an attempt should be made to decrease the 
number of surviving leukemic cells by purging. Dr. Cometta responded that the 
leukemic cells that are present do not necessarily contribute to relapse. Dr. Miller said 
that the protocol is specifically designed so that if relapse is shown to occur from cells in 
the marrow, then an attempt will be made to purge these cells from the marrow. 
Therefore, an attempt should be made to purge the marrow initially. 
Dr. Leventhal expressed concern that normal marrow cells will be transduced. Although 
the experimental design will provide information regarding the origin of relapse, no 
useful information will be obtained about improving the quality of purging. If the 
investigator demonstrates that relapse arises from the transplanted marrow cells, the next 
logical question is what will be the efficacy of purging. 
Dr. Mclvor stated that it is unclear how the investigators will determine if the leukemic 
cells that occur at relapse contain the marker gene and if the tumor cells can be 
distinguished from normal cells. Dr. Cometta explained that both bone marrow and 
peripheral blood samples will be obtained from the patient. Their cells will be isolated 
and grown in methycellulose culture, with and without G418. The morphology of the 
resulting colonies will be examined. PCR analysis will be performed on both G418- 
selected and nonselected colonies. In addition, these cells will be sorted by FACS. 
Dr. Leventhal noted that the protocol requires that the 2 x 10 6 cells for PCR analysis, 1 x 
10 7 cells for Western blot analysis, and 2 x 10 6 cells for methylcellulose culture and that 
these cells will be obtained from 5 milliliters of bone marrow. It is often difficult to 
acquire this number of cells from normal individuals. Dr. Leventhal suggested revising 
the protocol to require enough bone marrow to obtain sufficient cell numbers. 
Dr. Erickson had three concerns: (1) Is the frequency of transduction high enough to 
make this an evaluable study? (2) Can leukemic cells be separated from normal bone 
marrow cells, and can the contribution of marked cells to relapse be determined? and 
(3) Should a patient have to pay for a procedure that is not going to offer any 
therapeutic benefit? He said that he was satisfied with Dr. Cometta's response to the 
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