Protocol 
RAC Application 
Kenneth L. Brigham, M.D. 
Although cationic liposomes can deliver any nucleic acid construct, most work with this 
technology for in vivo delivery has concentrated on delivery of DNA in the form of plasmids. 
Plasmids associate readily with cationic liposomes by charge-charge interaction and, at least 
inlungcellsincultureand in vivo , cationic liposomes deliver functioning plasmids effectively. 
Plasmids do not cause human disease, are easy to work with in the laboratory and can be 
produced in large quantities. Although plasmids do not readily incorporate into host DNA and 
do not replicate in mammalian ceils, some investigators have found expression of transgenes 
imbedded in plasmids for several weeks following transfection (8). 
Rationale for alpha - 1 antitrypsin gene therapy An inherited deficiency of the principal 
endogenous antiprotease, alpha-1 antitrypsin (AAT), results in development of emphysema and 
respiratory failure at an early age (18). Although the exact incidence of abnormalities in the 
AAT gene is not known, it now appears certain that the disease is not rare. Current therapy 
consists of parenteral administration of the AAT protein at monthly intervals, although 
whether this treatment alters the course of the lung disease in these patients is unclear. 
Disadvantages of current therapy include expense, the necessity of repeated intravenous 
infusions and reactions to the protein either because of impure preparations or due to 
immunologic responses. 
We believe that if a functioning human AAT gene could be introduced into lung cells 
accessible through the airways, that the resultant local production of the antiprotease might 
be sufficient to protect the lungs f rom proteolytic injury, even though the total amount of AAT 
produced might be less than usually made in the liver. If the treatment is safe and efficacious, 
aerosol administration of plasmid/liposome complex at intervals probably in excess of a week 
would be trivial therapy compared to that currently available. 
In addition to the treatment of patients with AAT deficiency, we are interested in 
expanding the application of gene therapy to the treatment of acute lung injury. Acute lung 
injury resulting in the clinical entity called the adult respiratory distress syndrome (ARDS) 
occurs in approximately 1 50,000 Americans each year and mortality remains in excess of 50%. 
Knowledge accumulated over the past several years implicates the inflammatory response, 
particularly neutrophil activation, as important in the pathogenesis of lung injury. Neutrophils 
injure lung cells in part because of release of neutrophil elastase which is inactivated by AAT 
(19,20,21). In animal models and cultured cells, there is evidence that AAT can prevent injury 
of lung cells consequent to inflammation (21,22). Thus, transient hyperexpression of the AAT 
gene in the lungs of patients with or at risk for developing acute lung injury could be 
preventive or therapeutic. 
Because of our ultimate therapeutic goals, we propose to determine whether expression 
of an AAT transgene can be achieved using a plasmid/liposome delivery system in both 
patients with AAT deficiency and in patients without AAT deficiency. In the former case, we 
choose to begin with nasal instillation because there is already clinical experience with this 
approach, cells from the nasal mucosa can be easily obtained, no invasive procedures are 
necessary and AAT concentrations in nasal wash fluid can serve as a convenient and simple 
monitor of transgene expression. In the latter case, we propose to select patients who are to 
undergo elective lung resection, most commonly for cancer, and to instill the plasmid/liposome 
into the lung which is to be removed surgically. We believe that this approach has the dual 
advantage of minimizing any risk to the patient and of obtaining lung tissue which will permit 
more detailed assessment of transgene expression and toxic effects of plasmid/liposome 
delivery to the lower respiratory tract. We believe that this experimental design is conservative 
in that it minimizes risk and maximizes the information which can be obtained from the 
human studies. 
Recombinant DNA Research, Volume 19 
[177] 
