Recombinant DNA Advisory Committee - 12/3-4/92 
cells. No cytopathic effects were detected at an MOI less than 100 with purified vector; 
however, effects were observed at an MOI of 1,000. In conclusion, at the doses proposed 
for this study, host cell shut-off and cytopathic effects should not be problematic. 
In regard to the characterization of the production line, 7 preclinical production lots 
have been prepared. All of these lots meet the specified criteria. One of the 7 lots had 
3 colony forming unit (CFU) of bacteria. 
Dr. Welsh responded to Dr. D. Miller's concerns regarding wild-type virus production. 
Wild-type virus will be monitored by PCR and the HeLa cell infectivity assay. For the 
biological assay, 10 7 cells will be plated onto each of 10 plates. These cells will be 
subsequently infected with the vector at an MOI between 1 and 50. Using this criteria, 1 
wild-type virus particle should be detectable in 5 x 10 8 Ad virus particles. Therefore, 1 
wild-type virus particle should be detectable in the maximum administered dose. 
Dr. Welsh noted that the original protocol stated that the rat-1 cellular transformation 
assay will be used. Since Dr. Ginsberg and the RAC have noted that this assay is not 
particularly sensitive, Dr. Welsh asked that the requirement for this assay be deleted 
from the protocol. 
CF is a lethal genetic disease, and the proposed therapy could represent a major advance 
in the treatment of this disease. This protocol is designed to provide maximum safety to 
the participants by applying a small amount of virus to a defined region of the nasal 
epithelium. This study will provide critical data about biochemical efficacy and safety 
that allow for the design of new generations of vectors and the initiation of trials to 
assess clinical efficacy in the lung. 
In response to Dr. Carmen's question regarding the proposed vector, Dr. Welsh 
explained that this vector is based on a Type 2 adenovirus; Drs. Crystal and Wilson are 
using Type 5 adenoviruses. There is substantial sequence similarity between the various 
vectors. All 3 vectors have the El region deleted; however, there are different promoter 
regions. This vector has an Ad-2 Ela promoter, which is a moderate strength promoter. 
The other 2 CF studies will use high level expression promoters, either the major late 
promoter or a cytomegalovirus (CMV) enhancer beta-actin promoter. The 
polyadenylation sites are different between the proposed vectors. The Ad2/CFTR-1 
vector has an Ad-2 Elb polyadenylation site, the other 2 vectors will use SV40. The 
Ad2/CFTR-1 vector has retained the E3 region unlike the other vectors. 
There has been some discussion regarding E3 positive versus E3 negative vectors. This 
E3 positive vector is larger than the wild-type virus, thus its growth is impaired even in 
the permissive 293 cells. The large size of this vector offers an additional safety 
advantage. The wild-type adenovirus out competes the E3 adenovirus vector. Mixing 
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Recombinant DNA Research, Volume 16 
