Gene Therapy for CF using Cationic Liposome Mediated Gene Transfer: Phase 1 Trial 
bb. Nasal brushings and washings . Nasal mucosa will be pre-treated with 
0.05% oxymetazoline hydrochloride (Afrin) and topically anesthetized 
with 2% Lidocaine five minutes before brushing. Using a Pap smear 
cytology brush, the nasal mucosa at the site of gene administration below 
the inferior turbinate will be brushed gently to remove non-adherent cells. 
Cells recovered in the brushings will be disassociated in phosphate- 
buffered saline. Cystospin of these cells, as well of cells washed from the 
nasal mucosa with general application of a moistened cotton-tipped pledget 
will be analyzed for cell count, morphology, and cytopathic effects. 
b. Efficiency of gene transfer . 
1) Determination of gene transfer efficiency will be based predominately on 
measurements of the nasal potential difference. When ions move across the nasal 
mucosa, ion currents (and therefore potential differences) are generated. This 
potential difference is known to be significantly elevated in CF patients compared 
with non-CF controls. This well-characterized electrical abnormality of the CF 
nasal airway also reflects the abnormalities observed in the trachea and lower 
airways of CF patients. In order to evaluate nasal potential difference, a reference 
electrode (a 21-gauge butterfly needle filled with a 4% agar/Ringer’s lactate 
solution) is inserted subcutaneously in the volar aspect of the forearm. The 
reference bridge is connected through a calomel cell in 3M KCL to a voltmeter 
(BK Precision Instruments, Model 2906). An exploring bridge is established by 
inserting a second agar/Ringer’s lactate solution-filled needle into a PE 50 tubing 
line through which normal saline is being perfused at the rate of .2 cc per minute. 
The exploring bridge needle is connected through a calomel cell in 3M KCL to 
the volt meter. Nasal potential difference is measured by gently placing the tip of 
the exploring PE tubing line against the nasal mucosa, and can be used to 
evaluate different areas within the inferior turbinate, the septum, or elsewhere 
within the nose. Because the nasal PD is measured essentially instantaneously, 15- 
20 measurements at different points in both the experimental and control sides of 
the nose can be generated in a relatively short (10-15 minute) period. Measure- 
ments within the area of lipid/CFTR gene and control (no CFTR) administration, 
as well as other regions within the nose not receiving gene (e.g., septum, lateral 
wall) will be studied in both nares. The nasal potential difference can be 
performed repeatedly without damage to the nasal mucosa. Dr. Sorscher has 
previously performed this test on 10 CF patients and 15 non-CF controls without 
complications, and with verification of elevated potential difference in the CF 
patients. Correction of this elevated potential difference following gene 
administration (without change in the patient’s opposite nostril which receives no 
CFTR gene) would serve as evidence for gene transfer of the active CFTR. Nasal 
PD measurements within a given CF patient are quite stable; for example, daily 
PD measurements over a two-week period on one CF patient performed by Dr. 
Sorscher showed no statistically significant differences. Nasal PD measurements 
are currently being used as part of a study preliminary to CFTR gene transfer at 
The Children’s Hospital of Alabama, Division of Pulmonary Medicine, amiloride 
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Recombinant DNA Research, Volume 18 
