Bar Harbor, Maine, Despite years of work, high
hopes and some genetic finagling, scientists trying to use gene therapy
hemophilia - the bleeders' disease - face at least one more hurdle before they declare victory.
In pioneering experiments aimed at correcting
an inborn error - an inability to make a clotting factor that stops blood
from wounds - they have succeeded in dogs. But dogs aren't people, and the same treatments applied to men have worked
only temporarily. A permanent or long-term cure has remained disappointingly elusive.
The goal of such work, which has been under way
for more than a decade, is to correct the genetic error. Because patients
with hemophilia inherit a mutant gene, their bodies are unable to make one of the clotting factors needed to stop the bleeding
from wounds. As a result, even small cuts and bruises become medical emergencies.
As a rule, hemophilia is seen only in males, because
the gene is sex- linked, meaning it resides on the X chromosome. Men
have just one X chromosome, so they get no back-up gene in case one is faulty.
The experiments involve delivering new genes into
a patient's muscles or liver. The agent that transports them is a genetically
engineered virus, known as a vector. So far, the liver seems to be the best target, and genes inserted by the viruses worked
well - briefly - in one patient. Two earlier patients, who received virus injections into muscle, showed signs that the treatment
works, but at too low a level to do much good.
Geneticist Katherine High, who is running these hemophilia experiments
at the University of Pennsylvania, discussed her team's
latest results at a recent science journalists' meeting here at the Jackson Laboratory, a private research center that is a major
supplier of laboratory mice. She said exciting progress has been made - "we can do a great job in dogs" - but the big prize,
curing patients of hemophilia, is still beyond reach.
High said the latest patient tolerated the treatment
without side effects, and his liver accepted enough of the gene-laden viruses
to make effective amounts of the clotting factor. His tendency to bleed profusely when injured was reduced, but the benefits
"We got [gene] expression of about 12 percent"
of normal, which is enough to greatly ameliorate the disease symptoms,
said. But then the amount of Factor 9 found in his blood "went down to 11 percent a week later. And then to 10 percent, then
6 percent and 3 percent, and then it went all the way down," she said.
"When it was still at 10 percent we were pretty excited," High recalled.
"And then when it went all the way down again, I was
devastated." And, worse, "we haven't been able to figure it out."
David Curiel, a leading gene therapy researcher at the University of
Alabama, Birmingham, said, "I was really discouraged" by
that treatment failure. "That thing highlighted how animal results sometimes aren't completely transitional to human systems. "
Using a large animal model, Curiel added, "is always good because it
recapitulates many of the problems you can have with a
human. So her work in the dog was really excellent."
These dogs have a form of hemophilia that closely resembles the human condition.
The potential reasons for treatment failure in humans are numerous,
and High said her research team is gradually sorting through
them. A strong candidate is that something touched off an immune reaction that ended up killing the liver cells infected by the
gene-carrying virus, AAV. This reaction could have been spurred by the virus itself, with his body assuming the engineered
microbe was a dangerous infection. Or, High said, the patient's new supply of Factor 9 being made in the liver may be too
"foreign," also causing an immune reaction. Or an unknown contaminant may have been injected along with the engineered
viruses, similarly causing an immune response.
Nevertheless, High said, there is room for optimism: All three patients
treated so far have shown no dangerous side effects.
There are concerns that some viruses being used might accidentally cause diseases, even cancer. Such occurrences have been
reported in efforts to cure other genetic disorders.
The most recent problems arose in France, where two young boys among
nine treated for a genetic immune disorder were
discovered to have cancers, leukemia and lymphoma. Their doctors are virtually certain the so-called retroviruses that were
used to carry new genes into these patients were the culprits. In both instances, tests showed that the viruses had stitched
themselves into the patients' chromosomes in a way that stimulated excessive cellular growth, touching off cancer.
High said both young patients are still alive - in fact, the gene therapy
cured their genetic defect, which had caused a complete
lack of immunity - and were receiving treatment for their cancers. Both had been born with mutant genes that fail to make a
necessary enzyme, keeping them from mounting an immune response to fight infections. These are the kind of children who end
up living in isolation inside sterile plastic bubble chambers.
Even though such children can sometimes be rescued with bone marrow transplants, suitable donors can't always be found.
The experiments followed similar treatments developed at the National
Institutes of Health, in Bethesda, Md., done to provide
functioning immunity to children.
Despite the two cancer cases, the gene therapy used in France and elsewhere
has been successful. "Every kid who got new
genes is still alive, and that's a 100 percent success rate" in cases where the therapy "took," High said.
But problems also have developed with another common virus used as a "vector" to deliver new genes.
Almost four years ago, a young Arizona man, Jesse Gelsinger, died of
an immune reaction when researchers at Penn tried to
treat an inherited metabolic disorder, ornithine trans-carbamoylase deficiency. The disorder can lead to death, primarily in men.
Researchers modified a common virus, adenovirus, as the vehicle to get
copies of the healthy gene into Gelsinger. But his
immune reaction proved lethal. Gelsinger's death - plus the news of two French cancer cases - dealt blows to the field of gene
therapy. Many experiments were put on hold, and the use of retroviruses as vectors came to a halt.
In targeting hemophilia, High and her colleagues at Penn chose to employ
another virus, one that infects almost everyone during
childhood, the adeno-associated virus. It is not known to cause any disease or disease symptoms and is unable to reproduce
itself without help from another virus, such as adenovirus or herpes virus.
Still, the problems remain formidable. First, even if the gene can be
inserted into patients, it also has to work, preferably for a
lifetime. Second, it has to make enough of the missing protein, a clotting factor, to do some good. Fortunately, even a little does
a lot of good. Getting only 2 percent of the normal blood level of Factor 9 helps, and "5 percent gives a pretty normal life
without the risk of serious bleeding," High explained.
As for the virus, AAV, "it's a small virus, and most of us are infected
as children," she said. "And it has no known association
Most of the virus' own genes are removed and then replaced with the
gene that can make a blood-clotting factor. But the
researchers also had to ensure that the virus can still infect tissues where it is infused, in muscle or liver.
"We went into the hemophilia dog model," High said, where "we could
get long-term expression that was adequate" to alleviate
When the experiments began in patients, with injections into muscles,
"all of them showed good evidence for gene transfer and
expression, but the levels [of protein made] were too low to ameliorate the disease," High said.
There was evidence that infusing the doses into the liver would lead
to higher production of the blood-clotting factor. The data
from dog experiments was good, "we could see expression [of the inserted genes] for about five years," and the new gene
raised the protein level in blood to 5 percent or 10 percent of normal, "which changed them into dogs with mild hemophilia"
instead of severe hemophilia.
The experience in volunteer patients was encouraging and disappointing.
High said there was no toxicity from the treatment in
the first two low-dose levels but also little effect on the two men's hemophilia. At the third, higher, dose level that went into the
liver "we got expression of about 12 percent, then 11 percent, and the next week at 10 percent."
"And it looked, at four weeks out, like he had knocked out all of the
cells" that had been infused into the patient's liver. "We
have no answer why."
Is she discouraged? Yes, to some degree, High said. But the researchers
also had seen solid evidence that the treatment can
work, and at least that one patient "had touched the rainbow."