Human Gene Therapy Subcommittee - 7/30/90 
detectable at all. However if normal human PBLs were used, there 
was human IgG In the PBLs of the mice. However, there was 
individual variation in levels of IgG expression. 
Dr. Bordignon said that Southern gel analysis was run for the 
presence of human ADA in the spleens of the mice, and once again 
the mice treated with ADA-deficient human PBLs showed no ADA, 
whereas mice treated with ADA-positive PBLs showed the ADA band. 
In order to demonstrate the human ADA band, it was necessary to 
increase the concentration of the protein lysates by 4-10 times. 
Utilizing PCR analysis, he was able to see CD4''' T cell clones in 
the spleen. He said that his conclusion was these cells 
represent one single clone with one single integration event. He 
noted that the controls were IL-2 -stimulated PBLs which showed 
significantly more activity than can be expected when using the 
transduced cells. All clones were tested in parallel with PCR 
and were positive with the exception of one which was negative. 
Dr. Walters thanked Dr. Bordignon for making the long trip from 
Milan and for presenting the data. He then called for a brief 
recess before asking questions of Dr. Bordignon. 
Dr. Walters reconvened the subcommittee at 10:50 a.m., and asked 
Dr. Parkman to respond to Dr. Bordignon 's presentation regarding 
issues that had been clarified and to allow him to ask Dr. 
Bordignon specific questions. 
Dr. Parkman said he found the presentation clarifying in several 
regards : 
1. It was now clear that this was a series of experiments 
involving one patient, done over a period of months. 
2. It was clear there was a marked improvement in the 
child with PEG-ADA treatment. This leads to asking the 
question to what degree are the positive results in the 
mouse due to positive results in the patient and 
whether, if the patient had not responded as well, the 
mouse would have responded the same. 
3. It was observed that T cells from the patient's 
peripheral blood were capable of division in response 
to a specific antigen as a consequence of ADA therapy 
and that the child was able to make antibodies to 
tetanus toxoid. This means the T and B cells were 
capable of functioning in an antigen-specific manner 
prior to putting the ADA gene in. 
Recombinant DNA Research, Volume 14 
[71] 
