MJ. Welsh and A.E. Smith, RAC Application 
We have expressed human CFTR in transgenic mice under the control of the surfactant 
protein C (SPC) gene promoter and of the casein promoter (71,72). In those mice, CFTR 
was overexpressed in bronchiolar and alveolar epithelial cells and in the mammary glands, 
respectively. Yet despite the massive overexpression in the transgenic animals, there were 
no observable morphologic or functional abnormalities. (It is known that human CFTR 
expressed in recombinant mouse cells is functionally active, ref. 35.) In addition, 
expression of CFTR in the lungs of cotton rats produced no reported abnormalities (11). 
3.5 Use of Retroviruses for CF Gene Therapy 
Most gene therapy protocols that have been presented to the RAC have used recombinant 
retroviruses. Because of the previous scientific and regulatory experience with 
retroviruses, it seems appropriate to consider their use for gene therapy of CF. 
It is clear that retroviruses can be used to express CFTR in cultured epithelial cells; we 
have done so (35) as have others (23). However at the present time, several considerations 
would seem to preclude the use of retroviruses as a vector to treat CF. a) Most previous 
uses of retrovirus in gene therapy have treated cells ex vivo, but removal of target cells 
seems impractical for treatment of CF lung disease, b) Retroviruses require dividing cells, 
whereas airway epithelial cells divide infrequently, c) The airway epithelium is likely to 
contain 10 10 - 10 11 (73) epithelial cells. Even if it were possible to produce and deliver 
sufficient retrovirus vector to correct 1% of these cells in vivo ; a significant risk of 
insertional mutagenesis might be incurred. 
Thus, retroviruses do not currently appear to be the agent of choice for CF gene therapy. 
4.0 USE OF ADENOVIRUS FOR CF GENE THERAPY 
For our study, we propose to use a recombinant adenovirus as the vector to express CFTR. 
4.1 Differences Between Adenovirus and Retrovirus Vectors for Gene Therapy 
There are many differences between retroviruses (reviewed in 74), which have been used 
for most gene therapy protocols, and adenoviruses (reviewed in 75-77), the vector proposed 
here. Adenovirus contains a double stranded DNA genome of approximately 36 kb within 
a small icosahedral protein virion. The retrovirus genome is a single stranded RNA of 
approximately 7 kb, within a core surrounded by a lipid-containing envelope. Retroviruses 
infect cells by converting their genome into double-stranded DNA and integrating into the 
host cell genome; adenoviruses replicate extrachromosomally. Retrovirus gene expression 
is chronic and virus shedding continues over the life of the cell. Adenovirus infections lyse 
permissive host cells but may establish latent infections in others, e.g., lymphocytes and 
monocytes (78,79). Retroviruses contain minimally 3 genes encoding reverse transcriptase, 
gag proteins which comprise the protein core, and the envelope protein. Adenoviruses 
encode a great many more proteins including early proteins which are expressed prior to 
viral DNA synthesis and late proteins comprising predominantly virion structural proteins. 
Adenoviruses are stable to purification whereas retroviruses are not. 
Because the genome of the retroviruses is so simple, and because the viral gene products 
can be expressed without deleterious effect on the host cell, it is relatively straight forward 
to remove the endogenous retrovirus genes and to replace them with marker genes, 
antibiotic resistance genes or genes encoding therapeutic proteins (10,80). Such vectors 
can be grown in packaging cells that provide the missing gene functions to produce the 
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