Recombinant DMA Advisory Committee - 5/30-31/91 
the major points pertinent to the review of the protocol which were brought up at the HGTS 
and explain how those had been addressed in the materials and today's presentation. 
Dr. Mclvor said the first issue was safety. He noted that the same retrovirus (LNL6) was 
proposed to be used in this protocol had dready received FDA approval. However, he noted 
the major safety consideration related to the fact that hepatocytes were to be transduced 
rather than lymphocytes or bone marrow cells and that the risk of insertional mutagenesis was 
the major safety concern. He noted that the investigators had done an adequate job of 
estimating the probability of insertional mutagenesis in their supplemental materials and, 
despite there being a certain amount of unpredictable risk associated with this, it was not a 
major concern and that the investigators were aware of it. 
Dr. Mclvor said his second concern dealt with the actual migration of donor cells to the liver 
since this was a major issue in being able to detect tagged cells present in the liver and 
therefore a key factor in the feasibility of the proposed protocol. He noted that in a recent 
PNAS article by Ponder, et al, the indication was that this number of cells would be 1 in 
1,000. However, the baboon data provided in the supplemental material shows that it is 
possible to have as much as 5% of cells from the donor in the recipient organ. 
Dr. Mclvor said one of his concerns in the review for the HGTS meeting was the lack of 
information on transduction frequency and he said he felt the material presented this morning 
relative to this issue reassured him that PCR could be used to determine the presence of 
sequences with a sensitivity of 1 in 10^. He noted that one major question still remained in 
terms of transduction frequency and that was that the current experiments that were done with 
human hepatocytes needed to be scaled up to a level of 10^ hepatocytes to test the feasibility 
in humans. He invited Dr. Ledley to comment on this. 
Dr. Mclvor said he had questions also in terms of the evaluation of engraftment. His main 
concern was the application of PCR. He said he did not believe this was a major block to 
approving the protocol since Dr. Ledley had, in fact, provided some maps specifying exactly 
how the PCR would be done which demonstrated the sensitivity was at a level of 1 in l(r, 
which is a level necessary to detect the presence of the marker in the recipient liver. He said 
one other question he had was if one could perform in situ hybridization on liver samples to 
detect expression of the neo gene that this could possibly be a superior method of determining 
whether or not the marker was there. He asked Dr. Ledley to update the committee in terms 
of the development of in situ hybridization and confocal microscopy. 
Dr. Mclvor noted that he had asked Dr. Ledley to look into alternative methods of evaluation 
of engraftment since one of the major limitations in detecting donor cells by genetic tagging 
is that only a portion of the cells are actually tagged; these cells should be distinguishable on 
the basis of natural biologic polymorphisms. One method which Dr. Mclvor suggested needed 
to be looked at was the Y-specific protein or surface markers associated with the MHC, but 
he noted that these techniques had not yet been established for liver samples and therefore 
using these them would require major technical work-up before they could be assessed. Dr. 
Recombinant DNA Research, Volume 14 
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