M.J. Welsh and A.E. Smith, RAC Application 
Ad2-ORF6/PGK-CFTR Item 5 - Clinical Protocol 

BACKGROUND AND RATIONALE 
In our previously approved RAC and IRB protocol (Cystic Fibrosis Gene Transfer: In Vivo Safety and 
Efficacy in Nasal Epithelium) we provided a discussion of CF and CF lung disease as background and 
provided substantial references as documentation. We will not repeat that here. The major points are: 
a) Cystic fibrosis is a common genetic disease (1). 
b) Although there are a variety of clinical manifestations, severe chronic lung disease is the major cause 
of morbidity and mortality. 
c) Although improvements have been made, no treatment is directed at the underlying defect. As a result 
CF remains a life-threatening and often lethal disease. 
CONSIDERATIONS FOR GENE THERAPY OF CF 
In our previously approved IRB protocol (Cystic Fibrosis Gene Transfer: In Vivo Safety and Efficacy in 
Nasal Epithelium), we provided a substantial discussion of issues pertinent to gene therapy of CF and 
supporting references. We provide only a very brief summary here. A more extensive discussion is 
contained in Appendices 7.5 - 7.8 and in reference (1). 
Target Tissue for Gene Transfer. 
Because 95% of CF patients die of lung disease, the lung will be the main target for gene therapy. Within 
the lung, the target tissue for gene transfer that is most likely be of benefit to a patient with CF is the 
airway epithelium. Loss of function of CFTR within the airway epithelium is thought to contribute 
directly to the pathogenesis of the disease. 
It is unlikely that any gene therapy protocol will correct 100% of the cells that normally express CFTR. 
Our data in cultured cell models and that of other investigators suggests that correction of a fraction of the 
cells may be sufficient to at least partially correct the chloride transport defect (Appendix 7.7). Although 
the safety of overexpressing CFTR is not known with certainty, current data suggests that there may be a 
reasonably wide therapeutic index. 
Adenovirus Vectors for Gene Transfer to Patients with CF. 
Adenovirus has a number of advantages as a vector for gene transfer to the airway epithelium; these are 
enumerated in Appendices 7.5 - 7.8 and our previous RAC application. Adenoviruses also have an 
impressive safety profile in terms of the toxicity of wild-type infections, their use in vaccines, their lack of 
oncogenicity in humans, and our initial trials in humans. Adenovirus also has some potential 
disadvantages, the one of most concern relates to the possibility that administration of adenovirus may 
induce an inflammatory or immune response. An additional concern is the possibility of virus replication. 
Use of Nasal Epithelium to Assess Safety and Biochemical Efficacy. 
The use of nasal epithelium confers several advantages for our studies; we discussed these in detail in our 
previous RAC protocol and in Appendix 7.8. They include: a) The epithelium has the same morphology, 
and electrolyte transport as intrapulmonary airway epithelia. b) Upper and lower airway epithelia have the 
same defect in electrolyte transport in CF. c) The epithelium is accessible for vector administration, 
inspection, and sampling, thereby allowing relatively noninvasive evaluation, d) The risk to participants 
is minimized if an adverse reaction should occur, e) The effect of expressing CFTR Cl* channels in the 
apical membrane of the epithelium is readily assessed by measuring the in vivo transepithelial voltage (12) 
and its response to cAMP agonists, f) Any adverse effects of vector administration can be readily 
assessed by inspection, by sampling cells, cytokines, and inflammatory mediators, by assessing surface 
immunoglobulins, and by a full-thickness biopsy of mucosa. 
Evaluation of Clinical Efficacy of Gene Transfer in CF. 
A number of potential problems can be anticipated in the evaluation of clinical efficacy of gene transfer in 
the lung. As a result, it may be difficult to determine whether clinical benefits outweigh any adverse 
effects associated with vector administration. Because CF is a chronic disease, tests of severity made 
over a short period of time may not be sensitive or may be subject to error. If one chooses to study 
patients with severe disease (FEV1 of 20-30% predicted), then there will be a large amount of irreversible 
bronchiectasis and other pathology that would not be affected by gene transfer. Moreover, the two year 
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