They compared the ability of the four best-characterized jumping genes, or transposons, to insert themselves into a cell's DNA and produce a desired change, such as making the cell resistant to damage from radiation therapy.
They found the piggyBac transposon was five to 10 times better than the other circular pieces of DNA at making a home and a difference in several mammalian cell lines, including three human ones.
"If we want to add a therapeutic gene, we can put it within the transposon and use it to deliver the gene into the cell," says Dr. Joseph M. Kaminski, radiation oncologist at the Medical College of Georgia Cancer Center and a corresponding author on research published the week of Sept. 25 in the online Proceedings of the National Academy of Sciences Early Edition. "You can use these wherever retroviruses have been used."
In addition to piggyBac, researchers looked at what was believed to be the most efficient transposon in mammalian cells, hyperactive Sleeping Beauty, first found "asleep" in fish. They also looked at Tol2, another fish transposon, and Mos1, found in insects.
The piggyBac transposon, which has close relatives in the human genome, is widely used to genetically modify insects. Sleeping Beauty has been used to correct hereditary diseases, including hemophilia, in a mouse model.
For this study, researchers used transposons to deliver an antibiotic-resistant gene. "It's a way of screening and seeing which transposon is better," Dr. Kaminski says. They found that while piggyBac might not work as efficiently as a virus, it put Sleeping Beauty to shame when it came to making cells antibiotic-resistant.
"Sleeping Beauty has captured the field as far as transposons to be used in mammals," says Dr. Stefan Moisyadi, molecular biologist, at the University of Hawaii and a corresponding author. "But by comparing different transposons, we showed Sleeping Beauty is far inferior to piggyBac."
Scientists have used viruses as a gene delivery mechanism for more than 20 years because of their adeptness at getting inside cells and inserting themselves in DNA. But efficiency comes at a price. Gene therapy trials have been halted because of major complications, including deaths. As examples, one patient died because of his immune response to an adenovirus and three children in another study developed leukemia because the virus inserted itself upstream of a cancer-causing gene.
"With viruses, you don't have control," says Dr. Kaminski. "People have tried to modify viruses for site-specific integration and have not been very successful. Once they get into the cell, they can insert wherever they want."
Dr. Kaminski's previous work, published in 2002 in The FASEB Journal, hypothesized that the integration site for transposons can be selected. "Typically, viruses and transposons will integrate anywhere along the genome," he says. "If they integrate anywhere, it can obviously cause harm. If we can target the integration, be able to insert the gene at a safe spot in the genome, that would be beneficial." He confirmed that targeting integration is possible in a paper he co-authored in 2005 also in The FASEB Journal. "We can do it in insects," says Dr. Moisyadi. "I think it's a short step to take it to a targeting mechanism we can use in humans."
Another clear benefit is that transposons are cheaper to produce and probably safer than viruses. For example, retroviruses use RNA to make DNA, an error-prone process that must occur before integration, Dr. Kaminski says. Also, viruses can't carry larger genes, such as the dystrophin gene, which could help correct muscular dystrophy. On the other hand, unlike retroviruses, transposons have to be coated with lipid to slip into cells.
Although piggyBac is not as successful as the virus at integrating into DNA, "we could potentially make a hyperactive version of piggyBac, like they did for Sleeping Beauty, which might be as good or better than retroviruses," Dr. Kaminski says. "I think we'll do it or somebody will. I think it's a safer method."
"At the moment, unless something new comes out, it's the only way to go because viruses have been killing people," says Dr. Moisyadi, who has avoided viruses in his transgenesis studies.
"One of our next goals is to use transposons to deliver a radio-protective gene, called manganese superoxide dismutase, to potentially protect normal tissue from radiation damage," Dr. Kaminski says.
In cancer, he suspects gene therapy will focus on this type of modification of normal tissue for protective purposes as well as manipulating the immune response. However, it has broad applications for correcting single gene disorders, such as hemophilia, sickle cell disease and muscular dystrophy.
Toni Baker | EurekAlert!
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