By Janelle Weaver of Nature magazine

Scientists have developed a gene-repair kit that treats the blood-clotting disorder haemophilia in mice. The technique replaces genes in targeted organs without removing cells from the body, simultaneously correcting multiple mutations. It broadens the range of diseases that can be treated with gene therapy.

The method uses enzymes called zinc-finger nucleases. These are molecular scissors that replace specific DNA sequences by cutting through the double helix, after which the cell's repair machinery fixes the break.

Until now, therapies using zinc fingers have required cells to be taken out of the body, genetically modified in a dish and returned. This works for some immune and blood disorders such as sickle-cell anaemia, and trials are underway for HIV and diabetic neuropathy, but not for diseases affecting tissues less suited to this type of manipulation.

To develop a way to correct mutations within the body, Katherine High, a haemophilia researcher at the Children's Hospital of Philadelphia in Pennsylvania, teamed up with experts on zinc-finger nucleases at Sangamo BioSciences in Richmond, California. Their work is reported today in Nature.

Gene swap

People with haemophilia B have multiple mutations in the F9 gene, causing them to lack a blood-clotting factor made by the liver. As a result, their bodies are unable to staunch bleeding, and injuries leave them at risk of fatal blood loss.

To model the condition, High and her team used mice engineered to carry the faulty human gene. They designed zinc-finger nucleases to cleave the genome at the start of the F9 sequence and insert the unmutated gene, fixing all the mutations at once.

The researchers injected the mice with the zinc-finger nucleases, along with a liver-targeting virus modified to carry the normal version of F9. After treatment, the animals' blood clotted in 44 seconds, compared with more than a minute for mice with haemophilia, and contained 3-7% of the typical amount of the missing factor.

In humans, such levels would result in only mild bleeding. The factor was still produced after partial removal and regeneration of the liver, showing that the genome edits persist and are passed to daughter cells.

The mouse genome contains 20 sites besides the F9 gene that are most likely to be affected by these particular zinc-finger nucleases. But the treatment snipped DNA at only one of these, with no ill effects. There were no signs of toxicity or changes in liver function in the mice over an eight-month period.

Early days

"In theory, almost all genetic diseases could be amenable to this type of treatment," says Mark Kay, a gene-therapy researcher at Stanford University in California.

But, he adds, "there are still some technical hurdles that have to be overcome before this is going to be a wide-scale medical therapy". Questions remain, he says, about how to get the right amount of DNA to the right cells, the risk that zinc-finger nucleases will cut the wrong bit of DNA and the availability of suitable viruses to carry the genetic payload.

"It's still early days," agrees Paula Cannon, who studies zinc-finger nucleases at the University of Southern California in Los Angeles. "I'm cautiously optimistic that this won't be at all hazardous, but it's appropriate to make sure that these treatments are indeed as safe as we hope they're going to be."

This article is reproduced with permission from the magazine Nature. The article was first published on June 26, 2011.