volume of 50-100/xL. After remaining in contact with the nasal epithelium for 15 minutes, the 
nostril will be gently washed with normal saline, and the material aspirated. 
Dosages: Each patient group will receive defined doses of the AvlCF2 construct. Doses 
administered will range from 5 x 10 7 PFU to 5 x 10 8 PFU, and 5 x 10 9 PFU. 
Day -1: History and physical examination will be repeated. Nasal potential difference 
measurements will be made in the treated and untreated nostrils to assess the physiological 
correction of transepithelial electrical potential difference measurements. The treated area will 
be identified by endoscopy and compared to the other side. 
Day 0: History and physical examination will be repeated. Subjects will receive intravenous 
sedation and topical anesthesia of both nares (treated and non-treated) of the nose with 2% 
lidocaine. Epithelial cells will also be collected from the nose by brushing for cell culture. 
A sterile, disposable bronchial cytology brush will be used to gently brush the nasal mucosa 
at the location where the recombinant adenovirus was administered. After 10 seconds of 
brushing, the brush is removed and the cells shaken into 2 ml of sterile culture medium. The 
cell suspension is then kept on wet ice and placed in tissue culture using standard techniques. 
In situ hybridization, immunochemistry and chloride intracellular measurements will be used 
to assess expression of the transfected CFTR cDNA. 
Following the collection of cells from the nose, the fiberoptic bronchoscope will be 
introduced. The larynx will be anesthetized with topical 2% lidocaine, and then the patient will 
be bronchoscoped. PD will be measured in a defined area of the trachea using measurements 
from the carina to establish a specific location. BAL, brushing, and multiple biopsies (5) will 
be performed in the lobe that will be treated. The lobe will be selected based on its 
radiological appearances. We will try to select a lobe that is representative compared to the 
overall radiological appearance of the chest. The lobe should be dependent during the 
bronchoscopy to avoid dissemination of the virus to other lobes. All other things being equal, 
we will select the right lower lobe. Subjects will receive the AvlCF2 vector (10 10 , 10 11 , or 
10 12 PFU) in the bronchus in a total of 20 ml sterile fluid. At the end of the procedure, the 
trachea will be "painted" witij a small amount of the suspension, using small volumes and 
working from peripheral to proximal location. The equivalent lobe on the other side will be 
used as a control. 
Day 3: History and physical examination will be repeated. Nasal potential difference 
measurements will be repeated, and then subjects will be bronchoscoped again. PD will be 
measured in the trachea. BAL will be performed for detection of virus, viral DNA, elastase, 
cytokines, and cellular composition, the latter to assess inflammation. Brushings of the airways 
will be obtained using disposable bronchial cytology brushes. Bronchial biopsies will be 
obtained for histology, in situ hybridization of the CFTR mRNA, immunochemical and 
physiological detection of the wild type CFTR. The treated lobe and the control lobe will be 
studied. Chest x-ray, perfusion scans, CT scans, mucociliary clearance scans, oxygen 
saturation, and pulmonary function tests will be repeated to detect any improvement in the 
function of the treated lobe. CBC and differential, PT, PTT, serum chemistry, and urinalysis 
will be repeated. 
Day 7: History and physical examination will be repeated. Chest x-ray, scans, oxygen 
saturation, and pulmonary function tests will be repeated. Pharyngeal swabs and stool will be 
obtained for adenovirus culture using the permissive cell line (293) which supports the growth 
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Recombinant DNA Research, Volume 17 
