Researchers know that RDEB is caused by a mutation in a single gene, whose protein - type-VII collagen - migrates to the lower side of the skin cell where it acts as a glue to anchor that cell in place. An ideal gene therapy for this disease would insert a copy of the gene into the skin cells where it could produce normal type-VII collagen. The problem is that the gene is too large to work in most standard gene therapy techniques.
Khavari thought that Calos' technique would provide a way around the gene's large size. "I'm excited that this approach has the potential to make gene therapy work in this disease and in a lot of cases where gene therapy hasn't been working," Calos said.
Calos' technique hijacks a mechanism used by a bacteria-infecting virus (called a bacteriophage) to integrate its genes into bacteria. The bacteriophage makes a protein called integrase that inserts a gene into a specific DNA sequence on the bacteria DNA. It turns out that humans also have a version of that DNA sequence. When the researchers insert a copy of the therapeutic gene and a gene coding for integrase into a human cell, the integrase inserts the gene into the human sequence.
To test Calos' technique in RDEB, Susana Ortiz-Urda, PhD, a postdoctoral fellow in Khavari's lab, took skin samples from four children with RDEB. The samples included skin stem cells that replenish the outer skin layer as it sloughs off. She then inserted the integrase DNA along with the gene that treats RDEB. In the lab dish, these cells all made normal levels of type-VII collagen.
Once the researchers established that the skin stem cells contained working type-VII collagen, they transplanted those cells onto mice where they formed normal skin tissue. The altered skin cells con
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Contact: Michelle Brandt
mbrandt@stanford.edu
650-723-0272
Stanford University Medical Center
15-Sep-2002