M.J. Welsh and A.E. Smith, RAC Application 
predictable. Furthermore, clinical studies suggest that the clinical course is sufficiently 
predictable to allow for the meaningful assessment of future protocols aimed at delivering 
the gene to airway epithelia. 
A.l.c. Is the protocol designed to prevent all manifestations of the disease, to halt the 
progression of the disease after symptoms have begu n to appe ar, or to reverse 
manifestation of the disease in seriously ill victims ? 
The protocol is designed to test the safety, efficacy and dose of Ad2/CFTR-1. Because 
application will be limited to the nasal epithelium, it will not reverse manifestations of 
disease. 
A.l.d What alternative therapies exist? In what groups of patients are these therapies 
effective? What are their relative advantages and disadvantages as compared with 
the proposed gene therapy? 
We described standard therapies for CF in Section 2.2. At present there are no therapies 
designed to correct the fundamental defect. Participants will continue with all their current 
therapies during the course of the study. 
A.2. Transfer of DNA for Other Purposes. 
Not applicable. 
B. RESEARCH DESIGN. ANTICIPATED RISKS AND BENEFITS . 
B.l. Structure and characteristics of the biological system. 
Provide a full description of the methods and reagents to be employed for gene 
delivery and the rationale for their use. The following are specific points to be 
addressed; 
B.l.a. What is the structure of the cloned DNA that will be used? 
B.l.a.(l) Describe the gene (genomic or cDNA), the bacterial plasmid or phage vector, and 
the delivery vector (if any). Provide complete nucleotide sequence analysis or a 
detailed restriction enzyme map of the total construct. 
The cloned DNA to be used in the protocol was derived from Ad2 DNA and fragments of 
CFTR DNA. All of the DNA samples were originally obtained from ATCC and the 
construct assembled at Genzyme. The DNA construct comprises a full length copy of the 
Ad2 genome of approximately 37.5 kb, from which the early region 1 genes (present at the 
5' end of the viral DNA) have been deleted and replaced by the cDNA for CFTR. 
Specifically, nucleotides 546 to 3497 of Ad2 DNA are replaced with nucleotides 123-4622 
of the published CFTR sequence with 53 additional linker nucleotides. The topology of the 
5' end of the recombinant molecule is illustrated in Figure 2. The nucleotide sequence of 
the portion of Ad2 molecule that has been manipulated is given as Appendix 8. The 
remainder of the viral DNA is published in reference 106. 
The predicted CFTR transcript comprises a hybrid 5' untranslated region containing 46 
nucleotides of Ela upstream sequences, 47 bp of sequences derived from synthetic linkers 
and 10 nucleotides derived from the CFTR insert. The CFTR coding sequence comprises 
nucleotides, 603-5045 of the recombinant virus and 104-4546 of the hybrid Ela-CFTR-Elb 
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Recombinant DNA Research, Volume 16 
