Recombinant DNA Advisory Committee - 12/2-3/93 
Leventhal remarked that Dr. Schuening's semi-annual data report form included a 
statement that the reason for closure of the protocol on Hodgkin's disease was due to 
reported toxicides associated with IL-3. Dr. Schuening responded that a subsequent 
minor modification has been submitted in which permission is requested to replace IL-3 
with the less toxic fusion protein, GM-CSF/IL-3, for mobilization of CD34( + ) cells in 
the peripheral blood from bone marrow. Review of this minor modification is in 
progress. 
In response to Dr. Brinckerhoffs question about the preclinical studies, Dr. Schuening 
explained that he has performed numerous experiments (in vitro and in vivo) using the 
proposed vector. The experiments have included small animals, large animals, 
established cell lines, and normal human hematopoietic progenitor cells. Data 
demonstrates that the GC enzyme levels in normal cells are increased above endogenous 
levels. These levels of increase are even higher in fibroblasts obtained from Gaucher 
patients, e.g., 12- to 18-fold over nontransduced controls. With regard to long-term 
animal data, canine experiments have demonstrated expression of neo R out to 4 years 
following transplantation of transduced marrow cells. The inability to detect the co- 
transfected ADA gene in these animals was due to the high level of endogenous ADA 
activity. 
Responding to Dr. Carmen's question about preclinical canine safety data, Dr. Schuening 
said that two animals have received LgGC transduced marrow cells and have been 
observed for approximately 6 months. No evidence of toxicity has been observed. 
Dr. Miller stated that, as a collaborator on this protocol, he would respond to the RAC's 
questions about the retrovirus construct and packaging cell line. It is difficult to conduct 
comparison studies between investigators using different vector suppliers because of 
commercial considerations. Development and validation of vectors is extremely costly 
for the companies involved. The 3 GC vectors reviewed by the RAC are more similar 
than different. All of these vectors are all based on the Moloney murine leukemia virus 
(MoMuLV). The LgGC vector has a modified t-RNA binding site that eliminates the 
problem associated with expression by protein binding. Preliminary data indicate that 
this modified binding site allows the vector to express the gene insert better in fibroblasts 
than other vectors; however, there is no definitive data about this benefit in bone 
marrow expression to date. However, in vitro transduction of human bone marrow cells 
with the LgGC vector demonstrates increased GC activity, which is an indicator of gene 
expression. 
Dr. Tom Reynolds from Targeted Genetics Corporation, Seattle, Washington, explained 
the RCR testing procedures. The most likely mechanism for generating RCR is 
homologous recombination between the vector and the retroviral genes inserted in the 
packaging cells. The LgGC vector lacks the retroviral env gene, and the PG13 packaging 
cells were made by independent introduction of the gag-pol genes of MoMuLV and the 
env gene of gibbon ape leukemia virus (GALV) into the NIH3T3 TK‘ cells. These 
modifications were introduced to reduce the probability of generating RCR by 
recombination between the env genes of the vector and the packaging cells. Targeted 
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Recombinant DNA Research, Volume 18 
