Human Gene Therapy Subcommittee - 4/5/91 
were leukemic, and how this was determined. Dr. Deisseroth replied that cytogenetic 
analysis using a specific chromosome marker for CML, the Philadelphia chromosome, 
was utilized. In terms of the number of colonies for which PCR data exists, there are 27 
colonies and nine of them are positive for the neo gene. Dr. Mclvor asked if he had 
looked for BCR-Abl in addition to PCR for the neo gene. Dr. Deisseroth replied that 
he had not, but noted that methylcellulose colonies from unfractionated cells in patients 
who are 100 percent Philadelphia chromosome positive, turn out to be 100 percent 
Philadelphia chromosome positive. Dr. Mclvor asked if the investigators were confident 
that all the cells were leukemic cells, even though molecular analysis was not done on 
the colonies. Dr. Deisseroth said he was confident, but that he could do the molecular 
analysis on BCR-Abl. 
Dr. Mclvor said he wondered to what extent the committee would like to see results on 
the specific system that is proposed. He thought it would be worthwhile to see some 
results showing the molecular analysis. Dr. Deisseroth said he would be happy to 
provide the data. 
Dr. Mclvor asked now many marrow cells are given in a transplant. Dr. Deisseroth 
replied that the usual infusion is 2 X 10* cells per kilogram of the patient's weight. Dr. 
Mclvor stated that there are 1.2 X 10^ cells, that are going to be exposed to vector, and 
if there are 10^ cells per ml., this results in 10 liters of transplant volume. He asked how 
the volume could be reduced. 
Dr. Deisseroth said he intended to further reduce the number of cells with a monoclonal 
antibody separation which removes most of the mature cells, which are irrelevant for the 
experiment. Dr. Moen added that, together with Dr. Deisseroth, they had done a patient 
transplant using 10 liters. Dr. Mclvor asked about the frequency of transduction in those 
experiments. Dr. Moen replied that it was 21%, with the cell recovery being about 92% 
of the cells originally available in the supernatant. 
Dr. Deisseroth said that his center frequently fractionates cells to reduce the number of 
cells, with the goal of removing leukemic cells or removing T cells which cause graft 
versus host disease. It is a routine procedure to reduce the number of nucleated cells 
much below that harvested from the patient, by removing populations which do not 
contribute to hematopoietic recovery. 
Dr. Mclvor said it might be useful for the committee to see some data that demonstrates 
that the investigators can deal with these large scale manipulations. 
Dr. Anderson said the FDA requires that information; with his protocols he made the 
FDA material available to the committee. 
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Recombinant DNA Research, Volume 14 
