Gene Therapy for CF using Cationic Liposome Mediated Gene Transfer: Phase I Trial 
(100 micromolar), a drug known to block the elevated potential difference present 
in CF (and to bring the PD into a range similar to that observed in normal nasal 
epithelium following amiloride treatment) and terbutaline (100 micromolar) which 
stimulates normal CFTR will be superperfused onto the nasal mucosa in order to 
quantitate the proportion of nasal PD in patients attributable to amiloride or 
terbutaline sensitive transport. 
2) Efficacy of CFTR gene transfer will also be evaluated using nasal biopsy, 
brushings, and washings obtained following administration of lipid/DNA 
conjugates. CFTR protein production and localization at the apical membrane will 
be studied using immunocytochemistry and antibodies raised against CFTR. The 
goal in antibody localization experiments will be to determine whether an apically 
localized CFTR can be observed specifically in biopsies following wild-type 
CFTR administration. As noted above, for the Delta F508 CFTR homozygotes 
studied in this protocol, no apically targeted CFTR is anticipated since the 
mutation leads to maturational arrest of the mutant protein within the endoplasmic 
reticulum. Our laboratories have previously used CFTR antibodies to localize the 
CFTR protein to the apical membranes of cell sections (see Morris, et. al. . 
included in appendix). In addition, localization of CFTR to apical membranes of 
respiratory tissues has been reported previously by Dr. John Cohn (Duke 
University) using affinity purified antibody to the carboxyl terminal of CFTR. Dr. 
Cohn is currently collaborating with our group in localization experiments. 
Finally, we have recently developed a polyclonal antibody to the CFTR first 
nucleotide binding domain. In preliminary studies, this antibody recognizes the 
apical CFTR protein in tissues of normal mice, but not in mice in which the 
CFTR has been deleted. All these reagents will be utilized to optimize the 
chances of observing normal human CFTR expression in the apical membranes 
of human nasal epithelial cells transfected with the normal CFTR gene. 
Nasal epithelial cells recovered in biopsy brushings and washings will also be 
used to test for the presence of CFTR mRNA by reverse transcription PCR 
techniques. The approach for reverse transcribed PCR based detection of human 
CFTR has been used previously both for detection of human CFTR expression 
following gene transfer to murine lungs using cationic liposomes (14) and to 
detect transferred CFTR in very small numbers of cells obtained following 
bronchoscopic brushing in human patients (49). This technique will permit a 
highly sensitive measurement of normal CFTR expression in vivo , and coupled 
with PCR based detection of CFTR wild-type and mutant mRNA’s (Sorscher and 
Huang , Lancet, 337:1115-1118, 1991) will allow confirmation of wild-type CFTR 
in biopsy specimens taken from study patients. 
m.F. ALTERNATIVE TREATMENTS 
Standard cystic fibrosis therapy (chest physiotherapy, nutritional measures, antibiotics) 
as well as experimental therapies (rh DNase) are also available for the treatment of CF. If a 
patient decides not to participate in this study, it will not effect his/her care at the University of 
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