Recombinant DNA Advisory Committee- 9/12-13/94 
Dr. Crystal said that in his study with 9 human subjects, no replication-competent virus has 
been detected. This result was not obtained with the present modified vector with CMV 
promoter. Dr. H. Ginsberg asked how the viruses are assayed. Dr. Crystal said that 
secretions from patients are tested for adenovirus on 293 cells. Dr. H. Ginsberg said that 
proper sampling should be bronchial alveolar lavage. Ms. Meyers asked if there were any 
adverse effects on the 9 patients studied. Dr. Crystal said only the one that has been 
reported previously; after dose and inoculum volume reduction, no additional adverse 
effects have been observed. 
Dr. Samulski asked to compare the different adenovirus vectors used in all approved CF 
protocols, some are E3 plus, some are E3 deleted, and some are temperature-sensitive 
mutants besides the common El deletion. Dr. Crystal said that E3 expression required 
the presence of El; and in all the El deficient vectors, it was not relevant if the E3 was 
present or absent unless it was under the control of a constitutive promoter. Regarding 
the temperature-sensitive mutants, they are leaky; and they work in mice but not in 
humans. The other strategy is to delete E4 and E2b, but these vectors are still under 
development. 
Dr. Samulski asked about the stopping rule for the present study if an adverse effect is 
observed. Dr. Crystal said that if there was no safety issue, the study would continue. No 
virus shedding was observed when the vector was applied to the lung. 
Dr. Miller said that a published work reported vector replication observed in human 
epithelial cells reconstituted in nude mice. Dr. Crystal said that no vector replication has 
been seen in human studies. Dr. Miller asked questions about recombination with 
adenovirus sequences present in host cells. Dr. H. Ginsberg said it does not happen since 
293 cell sequences are integrated. Dr. Crystal stated that the criteria for clinical grade 
preparations are to assure that there is less than one replication-competent virus per 
patient dose. The clinical grade vector is prepared from a plaque-purified virus and 
treated with DNase to eliminate any contaminating adenoviral sequences. Dr. Miller said 
that the efforts to assure vector quality appeared adequate. Dr. H. Ginsberg said that his 
preparations used in animal experiments have not been as thoroughly prepared as the 
clinical grade materials. 
Mr. Capron asked if the patients were being treated with DNase. Dr. Crystal said that 70 
to 80% have proceeded through this kind of treatment. The patients will continue to 
receive DNase while on the study since it does not interfere with the present trial. 
Dr. H. Ginsberg asked if the cotton rat experiments have been performed with the 
original vector as well as the new CMV vector. Dr. Crystal answered that both of them 
have been tested. 
Dr. Smith asked if day 7 and day 30 are proper time points to look for inflammatory 
reaction with the E3 deleted vector. Dr. Crystal said the experiments have been 
conducted at both time points, and there is no difference. He will provide the data to the 
RAC. 
Recombinant DNA Research, Volume 20 
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