M.J. Welsh and A.E. Smith, RAC Application 
1.0 INTRODUCTION 
Cystic fibrosis (CF) is a common lethal, autosomal recessive disease of Caucasians (1-3) 
caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance 
regulator (CFTR) (4-6). CFTR is a chloride channel which is regulated by phosphorylation 
and by intracellular nucleotides (7). Mutations in the CFTR gene cause a loss of function 
of the CFTR chloride channel and thus, contribute to the hallmark of the disease: defective 
electrolyte transport by affected epithelia (1,8). Although there are a wide variety of 
clinical manifestations, lung disease, and particularly disease of the pulmonary airways, is 
the major cause of morbidity and mortality. Despite current standard therapy, the median 
age of survival is only 26 years. Unfortunately, no available therapy treats the basic defect. 
Gene therapy to deliver CFTR cDNA could correct the molecular defect; thus it would 
represent a major advance in treatment of CF. Because lung disease is currently the most 
severe clinical manifestation of the disease, the initial target cells will be those of the 
airway epithelia. The feasibility of gene therapy was initially demonstrated by our finding 
that expression of the cDNA for wild-type CFTR corrected the chloride c hann el defect in 
cultured CF airway epithelia (9). Our eventual goal is to deliver human CFTR cDNA to 
the airway epithelia of CF patients. For these studies, we will test a recombinant type 2 
adenovirus as a vector. Adenoviruses provide an attractive vector for introduction of the 
cDNA, because they appear to be safe, they do not require cell division for expression, they 
have a natural tropism for respiratory epithelia (10), and they have been used to deliver 
CFTR cDNA to the lungs of cotton rats (11). 
We have constructed a recombinant adenovirus vector in which the CFTR cDNA replaces 
the El region of a type 2 adenovirus, Ad2/CFTR-1. We have shown that this vector can 
transfer CFTR to airway epithelia in both cell culture models and in primate experiments 
and we have shown that it can complement the CF chloride transport defect. We have also 
assessed the safety of the vector system using in vitro cell culture models and in vivo 
experiments in hamsters and primates. 
We now propose to test the feasibility of using Ad2/CFTR-1 to deliver CFTR cDNA by 
applying the vector to a limited region of nasal epithelium in three CF patients. We will 
use the nasal epithelium for these studies for several reasons, a) Nasal epithelium has 
morphology and function similar to that of intrapulmonary airways, and nasal epithelia 
manifest the CF electrolyte transport defect, b) The nasal respiratory epithelium provides 
several advantages in safety for the first gene therapy trials because: the amount of virus 
can be minimized; access to nasal epithelia is easier and carries less risk than for pulmonary 
epithelia; and if adverse reactions should occur, the potential consequences for the patient 
are reduced, c) The use of nasal epithelium will allow us to assess noninvasively the 
efficacy of gene replacement by measuring directly the electrical potential difference across 
the nasal epithelium. 
The goal of this study will be to: a) assess the safety of the current recombinant adenoviral 
vector when applied to human airway epithelium in vivo', b) assess the efficacy in 
correcting the CF chloride transport defect in vivo', and c) assess the effect of dose of 
recombinant adenovirus on safety and efficacy. 
The results of these studies will be important in the development of future studies designed 
to deliver CFTR cDNA to the pulmonary airways and in the design of future generations of 
adenovirus vector. 
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Recombinant DNA Research, Volume 16 
