Scientists use gene therapy to correct sickle cell disease in mice

December 13, 2001

For the first time scientists have corrected sickle cell disease in mice using gene therapy, according to a study supported by the National Heart, Lung, and Blood Institute (NHLBI) of the National Institutes of Health and published in the December 14 issue of Science.

"Scientists have been working to accomplish this since the creation of an animal model for sickle cell disease several years ago. Although much more research is needed before human application, this is a significant achievement that brings us closer to human gene therapy for what is a very serious genetic blood disorder," said NHLBI Director Claude Lenfant, M.D.

Sickle cell disease affects about 1 in 500 African Americans and 1 in 1,000 Hispanic Americans. The disease is caused by a mutation in one of the two genes that determines the structure of hemoglobin, a critical molecule found in red blood cells. Hemoglobin transports oxygen from the lungs to other parts of the body. In patients with sickle cell disease, abnormal hemoglobin molecules stick to one another and form long, rod-like structures. These structures cause the red blood cells to become stiff - assuming a sickle shape. The sickled red cells pile up, causing blockages and damaging vital organs and tissue. In the study led by scientists at Harvard Medical School and the Massachusetts Institute of Technology, mice were bioengineered to contain a human gene that produces defective hemoglobin, causing sickle cell disease. The defect is an amino acid substitution on the so-called "beta" chain of amino acids that makes up part of the hemoglobin molecule. Since no single mouse model perfectly mimics human sickle cell disease, the scientists performed the experiment using two different mouse models. One mouse model contained only defective human hemoglobin and the other model contained a mixture of defective human hemoglobin and normal mouse hemoglobin.

Bone marrow containing the defective human beta-hemoglobin gene was removed from the bioengineered mice and genetically "corrected" by the addition of an anti-sickling human beta-hemoglobin gene. The new gene produces a beta chain of amino acids that when incorporated into the hemoglobin molecule gives rise to a modified normal hemoglobin molecule that prevents the sickling process.

After adding the anti-sickling gene, the corrected marrow was then transplanted into other mice with sickle cell disease whose bone marrow had been removed by radiation. Three months later, blood samples from the transplanted mice showed a high level of expression of the anti-sickling beta-hemoglobin gene, verified by identifying high levels of anti-sickling hemoglobin protein in the blood cells.

"Gene expression continued for at least 10 months in all mice in up to 99 percent of their circulating red blood cells. Up to 52 percent of the total hemoglobin incorporated the anti-sickling globin protein," said Dr. Philippe Leboulch, principal investigator of the study and assistant professor of medicine at Harvard Medical School and the Massachusetts Institute of Technology. Leboulch noted that gene expression above 15 percent is likely to have some therapeutic benefit in human patients. Further analysis of the structure of the transplanted mice's red blood cells showed a dramatic reduction in the number of irreversibly sickled cells. For one of the mouse models transplanted, no irreversibly sickled cells could be detected. These mice also had changes in the density of the transplanted red blood cells that "showed a clear shift towards normal," according to the scientists.

Two signs of sickle cell disease - enlarged spleen and a characteristic defect in urine concentration - were also corrected following the gene therapy.

The "lentiviral" vector used to deliver the therapeutic gene is based on human immunodeficiency virus (HIV). However, unlike the HIV virus, the vector is not capable of replicating or causing disease.

"The next step is to see how effective this vector is in larger animals more similar to humans. It will also be important to assess the safety of the vector when it is produced in large quantities - in particular with respect to its ability to replicate," said Greg Evans, Ph.D., a scientist with the Sickle Cell Disease Scientific Research Group of the Blood Diseases Program within NHLBI.

In addition to vector safety, another scientific issue to be addressed before human application is the toxicity of the regimen used to partially destroy the bone marrow of the transplant recipient before he or she receives the genetically corrected bone marrow.

"A number of research studies are underway to develop less toxic regimens which would still allow the new bone marrow to produce normal red blood cells for the long term," added Evans.

The first human application of gene therapy for sickle cell disease would be done with autologous transplantation. In this procedure, some of the patient's own bone marrow cells would be removed and genetically corrected. The remaining original marrow would be partially destroyed to "make room" for the genetically altered cells, which would then be returned to the patient. Currently, the only cure available for sickle cell disease is bone marrow transplantation. In this procedure, a sick patient (the recipient) is transplanted with bone marrow from a healthy, genetically compatible ("matched") sibling donor. However, only about 18 percent of children with sickle cell disease have a healthy, matched sibling donor.

For other children and adults with the disease, treatment includes transfusions, which correct anemia and prevent strokes, and pain-killing drugs. In addition, a drug called hydroxyurea is used in adults to reduce the frequency of painful crises and acute chest syndrome. The drug's use in children is still being studied.

Other collaborating scientists on this study were from the Albert Einstein College of Medicine, INSERM in France, and Genetix Pharmaceuticals.
NHLBI press releases and information on sickle cell disease can be found online at

NIH/National Heart, Lung and Blood Institute

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