CONTACT: JENNIFER BROWN
Iowa City IA 52242
(319) 335-9917; fax(319) 384-4638
Release: Feb. 19, 2002
UI study uses mini-proteins to repair cystic fibrosis defect in experimental
fibrosis (CF) is caused by genetic mutations in a gene that encodes the cystic
fibrosis transmembrane regulator (CFTR) protein. Normal CFTR protein forms
channels for chloride ions to leave cells. The controlled outflow of chloride
ions maintains cells' electrolyte and fluid balance and allows bacteria to
be cleared from the surface of airway cells. This action prevents infection,
which is a leading cause of death in individuals with CF.
University of Iowa researchers have shown that shortened versions of the
normal CFTR protein can function like the normal full-length protein in experimental
models of CF disease. These findings may point the way towards strategies
that could turn the potential of gene therapy for CF into a reality. The findings
are reported in the Proceedings of the National Academy of Sciences online
Gene therapy uses disabled viruses to deliver the correct version of a gene
to cells. A shortened, functional CFTR protein would be useful for gene therapy
of CF because it would allow the use of a particular disabled virus that is
proving to be a safe, efficient gene therapy vector.
"We would like to be able to use adeno-associated virus (AAV) as a
delivery vehicle or vector in gene therapy for CF," said Lynda S. Ostedgaard,
Ph.D., UI associate research scientist in internal medicine, and lead author
of the study. "This is a really nice vector. It can enter airway cells,
it has a track record with other genetic diseases and, to date, its safety
profile is encouraging."
However, despite its advantages as a gene therapy vector, AAV is too small
to accommodate the normal CFTR gene.
"Our goal is to make a short, functional CFTR gene with a view to making
it short enough to fit into AAV," said Ostedgaard.
The research team made a series of mini-CFTR genes that produce shortened
protein in airway cells. Specifically, they deleted parts of the CFTR protein
from a region called the R domain. The UI team used two model CF cell systems
to test the ability of the shortened proteins to act like normal, full-length
CFTR in airway cells. These experimental models are designed to mimic, as
closely as possible, airway cells in humans with CF.
"One model uses airway cells donated from individuals who have CF.
These are exactly the cells that a gene therapy treatment would target to
correct the faulty protein," Ostedgaard explained. "We often get
these cells from individuals with CF who undergo lung transplants. Without
those people being willing to make donations, we would never be able to do
these types of studies."
The second model used mice genetically engineered to lack the normal CFTR
Ostedgaard and her colleagues had previously discovered that portions of
the R domain could be deleted. However, they also found that a few specific
residues were required for function of the chloride channel.
"Based on our earlier work, we hypothesized that if you took out much
of the R domain but kept a few of the critical sites, the shortened proteins
should function properly," Ostedgaard said.
In both models, the researchers found that each mini-gene made protein with
all the properties and function of the normal full-length protein. In fact,
two of the shortened proteins transported chloride as well as normal CFTR.
"We proved that we could actually take out a fairly significant part
of the domain without affecting the protein's ability to transport chloride
ions and without affecting its ability to get to the right place in the cell
to start working," Ostedgaard said.
Ostedgaard added that it was satisfying and exciting when their hypotheses
were confirmed by the experiments. However, she indicated that while the results
were a step towards producing a working CFTR protein small enough to use in
AAV, the research had not yet reached that point. Additional studies are still
required to generate an AAV vector that could prove useful in CF gene therapy.
This type of research also leads to a better understanding of the structure
of this important CFTR protein, which could influence development of drug
therapies to treat CF.
In addition to Ostedgaard, the UI researchers involved in the study included
Michael J. Welsh, M.D., the Roy J. Carver Chair in Physiology and Biophysics,
professor of internal medicine and physiology and biophysics, and a Howard
Hughes Medical Institute investigator; Joseph Zabner, M.D., associate professor
of internal medicine; Christoph Randak, M.D., Ph.D., a postdoctoral fellow
in Welsh's lab; and Daniel W. Vermeer, Tatiana Rokhlina and Philip H. Karp
all research associates. Arlene A. Stecenko, M.D., associate professor of
medicine at Vanderbilt University also was part of the team.
The study was funded in part by grants from the National Heart, Lung and
Blood Institute (one of the National Institutes of Health) and the Cystic
Fibrosis Foundation (through the UI Center for Gene Therapy of Cystic Fibrosis
and Other Genetic Diseases).
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