Recombinant DNA Advisory Committee - 2/10-11/92 
Dr. Hirano asked if it was difficult to conduct experiments to determine ADA expression 
in the CD34( + ) cells. Dr. Dunbar stated that normal volunteers were used to select and 
enrich CD34(+) cells, perform neomycin resistance gene transduction experiments, and 
transduce with the ADA vector. It is unclear if increased levels of transduced ADA can 
be detected over the background of normal CD34( + ) cells that are not ADA deficient. 
One experiment has been conducted. In order to obtain sufficient numbers of cells to 
perform this experiment, patients need to be apheresed. The investigators do not want to 
expose an ADA patient to the increased risk of apheresis when the cells would not be 
given back to the patients. Small numbers of cells obtained from ADA patients have been 
transduced and pellets have been frozen. These cells could be assayed for ADA 
expression as compared to ADA levels prior to the treatment. 
Dr. Hirano asked if the investigators would want to know the expression level in the 
CD34(+) cells prior to transfusion. Dr. Dunbar stated that while it would be beneficial to 
have the data, the delay in obtaining this data would increase the risk to the patients. The 
investigators are not concerned about the expression level in CD34( + ) cells before they 
are differentiated. Dr. Hirano asked about growing CD34( + ) cells in culture. Dr. Dunbar 
said that CD34( + ) cells will be cultured in the same manner as with normal cells. Cells 
could be cultured for five or six weeks, plated in methyl cellulose, and myeloid colonies 
examined. Other investigators hold varying beliefs on whether this procedure is an 
acceptable model for stem cell differentiation. This procedure does not yield lymphoid 
colonies. A transduced population could also be placed into a Whitlock- Witte culture (a 
lymphoid-specific long-term culture system for examining B-cells) in order to observe long- 
term expression. Unless T lymphocytes are injected into an immune-deficient mouse, it is 
difficult to determine ADA expression. In addition, the experiment has to be conducted 
with cells obtained from patients with ADA deficiency. 
Dr. Brinckerhoff stated that the risk from the transduction was estimated to be 2xl0' 14 and 
asked if a risk could be assigned for the holes in the repertoire. Dr. Anderson responded 
that the patients were probably in the 70 to 80% normal range. The risk of getting a life 
threatening infection probably would be 0.1%, which is much higher than the risk from the 
protocol. However, the benefit could be profound; the patient's would be cured of their 
disease. 
Dr. Gellert asked how the first two patients could be classified healthy enough by the 
investigators to be allowed in public yet deemed to need this treatment immediately or be 
at mortal risk. If the patients were at such high risk, they should not be in public; or if 
they were not at that much risk, the protocol should be performed on a separate group of 
patients. Dr. Anderson apologized for implying that the patients were at mortal risk. 
However, there are holes in their immune repertoire which create a finite risk that the 
patients could be exposed to an antigen that they have no protection against. Therefore, 
Recombinant DNA Research, Volume 15 
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