Gene Therapy for CF using Cationic Liposome Mediated Gene Transfer: Phase I Trial 
human CFTR) indicate that native CFTR expression occurs within airway epithelial cells as well 
as the submucosal glands of the lung (47). Extensive biophysical, histopathologic, as well as 
cellular localization studies indicate that the pulmonary epithelial cells, and particularly cells 
lining the airways, are the appropriate target for initial CFTR gene transfer experiments. In 
addition, based upon experiments in which CF cells lacking the normal CFTR have been mixed 
with CFTR-corrected cells, it has been estimated that between 3-6% of cells in an epithelial 
layer must be corrected in order to begin to obtain normalization of the CF transport defect (48). 
This data provides an estimate of a lower limit of the proportion of cells within a monolayer 
which must be corrected in order to achieve at least some normalization of CFTR dependent ion 
transport. 
In analogy to previous gene transfer protocols for CF, the intent of this project will be 
the direct administration of CFTR to airway epithelium (in this case using lipid/DNA conjugates) 
in vivo . The predominant target cells will be the surface epithelial cells within the nasal airway. 
In CF, the nasal epithelial layer exhibits the same ion transport defects that are observed in the 
lower airways, and thus forms an ideal model system for study of potential therapies for CF lung 
disease in vivo . Potential therapies for cystic fibrosis using pharmacologic agents (amiloride, 
adenosine) as well as the approved gene transfer based strategies, have used nasal epithelium as 
a testing ground in the past (12,16-18). The nasal epithelium is easily accessible to 1) 
administration of therapeutic agents, 2) measurement of the CF bioelectric defect, and 3) 
biopsy/scrapings/washings, etc., to evaluate inflammation or other toxicity. In addition, use of 
the nasal airway allows potential therapies for CF lung disease to be tested without placing CF 
patients at the risks associated with administration of experimental agents to the severely 
compromised pulmonary airways already present within this patient population. 
The optimal level of expression of CFTR within individual respiratory target cells is not 
known. It is likely that endogenous levels of CFTR mRNA are quite low (on the order of 1-2 
mRNA molecules per cell [49]), and it is anticipated that viral or plasmid-based gene transfer 
may result in much higher expression levels than those found endogenously. No CFTR related 
toxicity has been observed in CF or other cells in culture which over-express the CFTR protein. 
In addition, studies in transgenic mice in which human CFTR is markedly over-expressed in 
pulmonary epithelial (and other) cells have not indicated any toxicity attributable to production 
of the human CFTR (50). There is a theoretical possibility that expression of the wild-type 
human CFTR in CF patients could lead to development of anti-CFTR antibodies and other 
inflammatory responses directed against cells expressing the wild-type protein. For example, if 
the common Delta F508 CFTR is not normally present on the cell surface in CF patients, it is 
possible that wild-type CFTR in a homozygous Delta F508 patient could produce an immune 
response specific for human CFTR. However, this has not been reported either in transgenic 
mice expressing human CFTR, or in CF mice or rats expressing CFTR following lipid mediated 
gene transfer (11,50, also see below). 
I.B.7. Cationic Lipid Formulations for Use in Gene Transfer 
Several cationic lipid formulations can be used as gene transfer vectors to deliver plasmid 
DNA in vitro , although certain of these appear to show considerably enhanced efficacy in 
reporter gene assays in certain cell types. Feigner, et al .. (51) described the use of a 1:1 molar 
mixture of DOTMA with DOPE (lipofectin) to deliver a plasmid DNA containing a chloram- 
phenicol acetyl transferase reporter gene construct to COS-7 simian kidney cells. This 
Recombinant DNA Research, Volume 18 
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