Hemophilia: Finding the Right Path

For this article, Jim Kling interviewed Mark Kay, a professor of pediatrics and genetics, and director of the Human Gene Therapy program at Stanford University, Stanford, Calif., and pediatrics professor Katherine High, University of Pennsylvania and the director of research, hematology division at the Children's Hospital of Philadelphia. Data from the Web of Science (ISI, Philadelphia) show that Hot Papers are cited 50 to 100 times more often than the average paper of the same type and age. R.W. Herzog, E.Y. Yang, L.B. Couto, J.N. Hagstrom, D. Elwell, P.A. Fields, M. Burton, D.A. Bellinger, M.S. Read, K.M. Brinkhous, G.M. Podsakoff, T.C. Nichols, G.J. Kurtzman, K.A. High, "Long-term correction of canine hemophilia B by gene transfer of blood coagulation factor IX mediated by adeno-associated viral vector," Nature Medicine, 5[1]: 56-63, January 1999. (Cited in 110 papers) R.O. Snyder, C. Miao, L. Meuse, J. Tubb, B.A. Donahue, H.F. Lin, D.W. Stafford, S. Patel, A.R. Thompson, T. Nichols, M.S. Read, D.A. Bellinger, K.M. Brinkhous, M.A. Kay, "Correction of hemophilia B in canine and murine models using recombinant adeno-associated viral vectors," Nature Medicine, 5[1]:64-70, January 1999. (Cited in 95 papers) Little doubt exists that hemophilia is a good candidate for gene therapy.1 There isn't much disagreement over the choice of vector, either; the adeno-associated virus is considered relatively safe. Less certain, however, is the method of administering the vector. Hemophilia results from a mutation in a single gene that is involved in blood clotting (factor VIII in the case of hemophilia A and factor IX for hemophilia B). The proteins travel systemically, so genes involved in their production can be placed in any tissue that can export it to the bloodstream. The proteins are already used to treat the disease, which affects one in every 4,000 live male births, according to Hemophilia Health Services. In these two Hot Papers, teams led by Mark Kay, a professor of pediatrics and genetics and director of the Human Gene Therapy program at Stanford University, Stanford, Calif.; and pediatrics professor Katherine High, University of Pennsylvania, chose two similar vectors, but different pathways, and achieved comparable results. Kay and his colleagues chose to deliver vector to the liver, while High and her colleagues, following a long tradition, delivered genes intramuscularly. Courtesy of Childrens Hospital of PennsylvaniaKatherine High "If you look at the papers, you see that the maximum levels of clotting factor achieved are the same using either muscle or liver as the target. However, you have to use higher doses [of the intramuscularly injected vector]," says High, who is also director of research, hematology division at Children's Hospital of Philadelphia. That difference could be attributable to differences in the preparation of the vector, she adds. Injecting into the Liver In his paper, Kay and his colleagues report the correction of hemophilia B in mice and dogs by delivering factor IX through AAV in a much less invasive manner than previously conducted. This time, they injected factor IX into a portal vein entering the liver. The AAV had been stripped of its viral genome, leaving only its capsid. Unlike many of the earlier vectors, wild type AAV, from which the vector is derived, is not known to cause any human disease, making it safer than some alternatives. Kay spent years developing gene therapy protocols to treat hemophilia B. In 1993, his group reported on a partial correction of the bleeding diathesis in hemophilia B dogs using a retroviral vector.2 The only problem was the team had to remove two-thirds of the liver to do it. "No one would want to do that in the clinic," says Kay. Kay focused on delivering factor IX to the liver because that is the site of its production in healthy individuals. The transduction need not be overly efficient: a mere 5 percent of liver cells churning out factor IX can do the job because those cells can over-produce the protein, resulting in therapeutic blood concentrations. "We're up to near normal levels [of factor IX], and you probably need only 20-25 percent to essentially correct [the disease]," says Kay. In part, the work was a big step forward because it involved dogs with hemophilia B. These are good human disease models because a dog is a large animal and, genetically, are far-removed from mice, which do not always accurately predict responses in humans. "If you can treat the disease in two animals that are far apart and one is a large animal, it makes one more confident that the therapy is likely to do the same thing in humans," Kay says. His paper, he says, "goes on to prove the concept that you could treat hemophilia in a clinically relevant manner that was safe; there was no toxicity and we didn't have to do fancy manipulations." Injecting Intramuscularly High believes that intramuscular injection could have some important advantages. For one thing, many patients who would qualify for this treatment already have damaged livers, because many hemophiliacs contract hepatitis from plasma transfusions. "So we have an interest in developing a treatment strategy that could be used by as many people as possible," she says. "Most people have intact skeletal muscle whether or not they've been infected with hepatitis." Vector injection into skeletal muscle is also a simple procedure that could be easily done outside of a standard clinical setting. "Delivering it to the liver is more complicated, though I'm convinced that can be done safely, she says. "But it's appealing to have something you can just give as a shot.... It is estimated that over half of the world's hemophilia population doesn't have access to [standard treatments]. The possibility of a procedure that would be relatively simple, that you could do with access to sterile needles and alcohol, is appealing." The jury is still out on which method works best. The two groups have begun to collaborate since the publication of these papers, and they recently completed initial human trials with intramuscular injection. In eight subjects with three different doses, they found good evidence for gene transfer and expression. "So the animal models were good predictors of what happened in humans," says High.3 Still, they didn't reach the factor IX target levels that they accomplished in dogs, and they didn't witness consistent elevation to levels greater than 1 percent of normal, which is the minimum required to ease the clinical effects of hemophilia. But those patients who did achieve intermittent levels higher than 1 percent have gone on to require less frequent factor IX treatments, which suggests that the gene therapy may have garnered some clinical effect, says High. With that work completed, the two groups are about to start a Food and Drug Administration-approved human trial using liver delivery, though High anticipates they might return to intramuscular work as they refine their methods. "My feeling is that it would be ideal to develop approaches for both muscle and liver, and that way there will be an option if patients have hepatitis and can't take the liver approach," High says. Kay's group, which is also working on hepatitis, continues to investigate other viral and nonviral vectors, as well as strategies to increase the efficiency of transduction and ways of increasing the AAV's capacity. Jim Kling (jkling@nasw.org)is a freelance writer based in Washington, DC. References 1. P. Smaglik, "Gene therapy crossroads," The Scientist, 13[14]:1, July 5, 1999. 2. M.A. Kay, et al., "In vivo gene therapy of hemophilia B: sustained partial correction in factor-IX-deficient dogs" Science, 262:117-9, 1993. 3. M.A. Kay, et al., "Evidence for gene transfer and expression of factor IX in haemophilia B patients treated with an AAV vector," Nature Genetics, 24[3]:257-61, 2000.

Hemophilia: Finding the Right Path

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