Just as the discovery of DNA turned 50, a gene therapy clinical trial in France became an example of what can be accomplished, as well as what might go wrong, when genetic material is used as medicine.
Earlier this year, several children with X-linked severe combined immunodeficiency disorder (X-SCID) developed leukemia after receiving treatment with gene therapy. Although effective, the treatment has not been used again after French officials halted the trial. In the United States, gene therapy trials that also use retroviral vectors were halted for a time in late January, and then restricted.
Now, a study from researchers at the National Human Genome Research Institute (NHGRI) may help explain how the vectors used play a role in cancer development in treated patients. They found that one popular retroviral vector, the murine leukemia virus (MLV), tends to insert genes into transcription "start regions" of the human genome. Those are areas that can either turn adjacent genes on or off, increasing the likelihood that growth-promoting genes are activated and, in theory, lead to cancer.
Many researchers have been studying the safety of the vectors, hoping that development of leukemia in the children was a highly random and remote event. But after mapping hundreds of MLV integrations in a human cell line, the NHGRI researchers found the MLV vector "prefers to land" in and around promoters that activate genes.
"The data are troubling and there has to be some concern now about use of MLV, one of the most popular delivery vector vehicles at the moment, and the only one that has worked," said Shawn Burgess, Ph.D., head of NHGRIs Developmental Genomics Section and author on the study, which was published in June in Science. "We should start looking for viral vectors that have a safer profile."
The First Successful Gene Therapy
The French X-SCID trial "is probably the first cleanly effective gene therapy study, the first to show therapeutic use of a retrovirus vector," said Theodore Friedmann, M.D., the outgoing chair of the Recombinant DNA Advisory Committee (RAC) of the National Institutes of Health. "It is also the first one that studies the nature of the harm that is intimately connected with this effective technology, so it raises an interesting set of problems. We have now gotten to the point where we can look at real benefits and real risks."
|
The first T lymphocytes were detected in the childrens blood within several months, and production increased to normal levels within 6 months, protecting the children against bacterial and viral infections. The treated stem cells divide infinitely, passing the corrected gene to daughter cells.
But within 3 years, two of the boys developed T-cell acute lymphocyte leukemia (ALL), apparently because the MLV vector inserted its therapeutic gene on top of the oncogene LMO2, which has been shown to cause childhood leukemia.
Gene therapy is designed to correct a genetic defect by inserting a healthy gene into cells, and in the case of the French trial, "it was too effective, in a way," said molecular biologist Christof von Kalle, M.D., of the Cincinnati Childrens Hospital Medical Center.
|
But the possibility that leukemia could develop in patients was discussed as far back as 1995. "We already knew that if you insert retroviruses in critical places in the mouse chromosome you get ALL," said Stuart Orkin, M.D., a professor of pediatrics at Harvard Medical School, Boston, who co-chaired a report to the National Institutes of Health on gene therapy. "The issue in humans is how many hits one can get in a cell of chance integration near a deleterious gene, and most safety studies hadnt hit enough genes. The insertions next to LMO2 were obviously beyond chance."
Cancer in the children was successfully treated, but fallout from the incident was widespread. Most European countries put gene therapy experiments on hold, and in January, the U.S. Food and Drug Administrations Biological Response Modifiers Advisory Committee (BRMAC) suspended 27 experiments that were using vectors or genes similar to those used in the French trial. In late February, the advisory committee allowed some of the halted gene therapy trials to resume under close monitoring, but only if no alternative therapies existed for life-threatening disorders. The RAC took a different approach from the FDA, Friedmann said. "We felt the chances of doing good are greater than the documented risks with vectors."
But research pinpointing just how MLV acts within a cells chromosome became possible after the genome was sequenced, and in the Science study, Burgess and Xiaolin Wu, Ph.D., a postdoctoral researcher, repeatedly infected human cells with MLV and watched where the vectors went. They found that of 903 integrations, the virus hit a gene 34% of the timemuch more frequently than would be randomly expectedand that it was preferentially inserted into "the most important area of the gene, the 5-prime end where the promoters, enhancers, and regulator elements are located," said Wu.
"What we saw leads us to believe that in theory, all of the children in the French trial had many integrations around the LMO2 gene," said Burgess. "Even though this is the only study that has seen cancer as a side effect, it is the only trial that has worked, and so it raises a red flag."
Kalle cautioned that Burgess cell line study should not be overinterpreted. "It seems remarkable that the use of retrovirus vectors has rarely led to side effects compared to the level of risk [calculated by Burgess and Wu]," he said. "I assume this will remain a very interesting field of study for some time."
Future of MLV and Other Vectors
Although lessons can be learned about MLV, they may not apply across the gene therapy landscape, some researchers say. A major study of another commonly used vector, the adeno-associated virus (AAV), also found that it is preferentially inserted within genes, but researchers say their risk of causing cancer is probably much lower than MLV vectors.
The study, published June 1 in Nature Genetics, was led by Mark Kay, M.D., Ph.D., professor of genetics and pediatrics at Stanford University Medical Center, Palo Alto, Calif., who uses AAV vectors to deliver a clotting factor gene to patients with hemophilia B. The vector is also being used to treat cystic fibrosis. Given lingering questions about the safety of retroviruses, Kay tracked where AAV vectors integrated in the liver cells in mice and found that 72% of the time, the vector had interrupted a genea much greater rate than would have been expected if insertion was random.
Still, Kay maintains that AAV and MLV vectors are "like apples and oranges." The MLV vector used to treat X-SCID had the potential to enhance cell growth and were engineered to treat stem cells from the bone marrow. Trials involving stem cells are considered more risky because those cells proliferate, and leukemia is a disease in which blood cells proliferate, he said.
"In general, the AAV vector doesnt insert very efficiently, unlike the murine vector," Kay said, adding that while the potential to cause cancer with AAV is "still there, the likelihood is probably small."
But Abbey Meyers disagrees with the optimistic viewpoint shown by researchers who think the risks of gene therapy are either small, or manageable. "We knew from day one of the first experiment that the virus causes cancer in animals," said Meyers, president of the patient advocacy group the National Organization for Rare Disorders, and a critic of the way gene therapy trials have been conducted. "Explanation of that risk has always been required to be in the consent documents of every gene therapy trial, but many investigators just ignore those risks. ... I think we need to go back to do more animal experimentation because it is clear that the vector caused cancer, and its likely it can happen again and again."
But in the absence of a clear answer, the risks and benefits of gene therapy must be weighed on a patient-by-patient basis, many researchers say. "This is the only vector that has worked, and so you have to do a cost-benefit analysis, and make tough calls," said Burgess. "But its clear the prognosis for leukemia is better than for X-SCID." Wu added that although researchers are working to improve the design of the vector, "it will continue to be used because it is the best choice."
At a RAC meeting held in mid-June, the committee discussed trials with MLV vectors that have been allowed to proceed, and agreed that "the risk, as currently known, is acceptable in a narrow set of cases," said RAC member, bioethicist Madison Powers, J.D., D.Phil., director of the Kennedy Institute of Ethics at Georgetown University, Washington, D.C. "Several committee members drew the analogy to similar judgments of acceptable risk in many cancer treatments for which the risk of treatment itself is production of another cancer later.
"In my view, these are truly tragic choices, and the instances in which the risk would be acceptable are quite limited," Powers said. "Moreover, a change in our knowledge relevant to that calculation could change that conclusion about risk and benefit."
"There is no ideal vector for any particular application, no single vector that solves all the technical problems, so I dont think it is appropriate to walk away from retroviruses as a class of vectors," said the RACs Friedmann. "All of our current armamentarium and methods of delivery still need to be evaluated and developed and studied because they all have potential benefits and hazards."
![]() |
||||
|
Oxford University Press Privacy Policy and Legal Statement |