Editor's Note: In November of 2009 a French research team reported an amazing breakthrough. They had stopped the frightening progress of a fatal brain disease in two young boys in Spain by using a novel type of gene therapy. Not readily apparent in ensuing international headlines was the Salk Institute discovery that made this gene therapy possible. Here is the story behind the story.
It was the early 1990s and Salk Professor Inder Verma had the deadly Human Immunodeficiency Virus (HIV) in his sights. One of his institute colleagues had contracted AIDS after receiving infected blood during heart bypass surgery, and Verma witnessed the terrible efficiency of the virus and the dreadful consequences of the disease.
The HIV was proving a fierce adversary to researchers the world over. During the previous few decades, biologists had developed a basic understanding of how viruses like HIV work. A virus inserts itself into or otherwise hijacks our cells, using our cellular machinery to rapidly reproduce. Most viruses prey on our dividing cells, happily multiplying until our healthy immune systems prevail. But one of the HIV's most fearsome aspects is that it can also introduce itself into the genes of our non-dividing cells, like the cells in our brains and the immune system.
Verma, the American Cancer Society professor of Molecular Biology in the Salk's Laboratory of Genetics, started thinking about this unusual characteristic of the HIV in a rather radical and unexpected way. What if, he thought, "we could engineer the virus, entirely removing its ability to cause disease, while retaining its ability to enter non-dividing cells?"
Verma was already deeply engaged in the emerging field of gene therapy, and had an impressive track record of achievement. About 10 years prior Verma and a UCSD scientist successfully repaired a human genetic defect in animal cells in a Petri dish, making front-page news in The New York Times. He knew that there were limits in the choice of viruses that could be used to therapeutically deliver healthy genes (known as viral vectors). The available vectors could only effectively deposit their cargo into (correct the genetic defect in) dividing cells.
Here was this dangerous, yet intriguing HIV from the lentivirus family. Could Verma and his colleagues strip the HIV of its powers to cause AIDS, making it possible to use the defanged lentivirus to target areas like the nerve cells in the brain, or the rare population of stem cells that make our blood and the immune system?
The Salk Institute had just established an infectious disease laboratory working on AIDS, and one of the first faculty to join this group was Didier Trono, who knew how to grow the AIDS virus in the laboratory setting. Verma had recently taken on a young Italian post-doc named Luigi Naldini who was inclined to pursue this potential gene therapy tool. Moreover, Professor Rusty Gage had recently joined the Salk faculty. Gage had valuable expertise regarding mice and their nervous systems, and was a generous collaborator. A young German MD, Ulrike Bloomer from Gage's laboratory, was assigned to collaborate with Naldini.
In a recent interview with The New York Times, Naldini, who is now director of the Institute for Gene Therapy at Vita-Salute San Raffaele University in Milan, Italy, recalled his days at the Salk working on the HIV as a potential viral vector. "We were scared of course," he admitted. But Naldini knew that if he could remove enough of the genetic material that causes AIDS, he could render the virus harmless while preserving its key transportation function.
Verma and his team developed a way to incapacitate the virus so completely that all it could do was convey the genes. Their April 12, 1996 paper published in Science offered this concluding sentence: "We believe that the generation of safe and efficacious lentiviral vectors will significantly advance the prospects of human gene therapy." The paper made quite a stir. It was the scientific version of "beating swords into plowshares," Verma said, chuckling at the memory.
"Next we had to show that the virus could carry a foreign gene and had the ability to introduce that gene into other non-dividing cells and tissues in mice," he said. The team performed its exacting work, yielding a second Science paper in 1997, demonstrating that they could actually use this new viral vector to deposit a gene into bone marrow in mice and that gene would now be appropriately expressed. "We made the vector completely safe and showed it could be used to treat genetic diseases," he said.
After years of creative and intensive work, the Salk team had made a seminal basic science contribution to the gene therapy field – conceiving a novel use for the deadly HIV and proving, in animal models, that it could be harnessed to convey healthy genes. It was time for others to carry the ball toward potential human use of this new vector.
Verma and the Salk Institute patented the discovery in 1996 and then licensed it to a Bay-area biotech start-up company called Somatix Therapeutics (Verma and Gage were both co-founders) which later merged with Cell Genesys. The company's role was to develop gene therapy tools like his lentiviral vector and it seemed a reasonable match.
Michael T. White, senior director of the Salk's Office of Technology Management and Development, says it was a genuine breakthrough to have devised a viable delivery system for ferrying healthy genes into non-dividing cells, and the company could have pursued its development. But the merged company wanted to concentrate on what its leaders perceived to be the best opportunities for maximum sales at that time. So Cell Genesys initially focused on gene therapy systems for cancer and "back-burnered the lenti system," White said.
Meanwhile, toward the end of the decade, the entire field of gene therapy was buffeted by some dramatic, high-visibility setbacks. Most notable was the death of 18-year-old Jesse Gelsinger, the first person to die in a gene therapy clinical trial. Gelsinger suffered from a form of genetic liver disease that can be fatal in infants. He joined a clinical trial at the University of Pennsylvania and was treated with healthy genetic material carried by an adenoviral vector (a DNA virus vector) in an experiment intended to demonstrate the safety of the procedure. He had a massive immune response to the viral vector, triggering multiple organ failure. Gelsinger died four days after the therapy, in September of 1999.
(Verma co-chaired the NIH-RAC Committee to investigate the circumstances and causes of this unfortunate development, testified at a U.S. Senate hearing and made recommendations to reduce the likelhood of recurrence of this tragedy.)
Shortly after Gelsinger's death, in 2000 in France, a separate clinical gene therapy trial began to treat a form of SCID (Severe Combined Immunodeficiency), commonly known as Bubble Boy Syndrome. SCID is a genetic disorder that renders its victims extremely vulnerable to infections. Gene therapy improved the immune systems in 18 of 20 patients treated, and was labeled a success. But the trials were halted when it was discovered that the gene-carrying retroviral vector (a distant cousin of the lentiviral vector) was depositing its material in the wrong place in the human genome, inadvertently turning on an oncogene. Five of the children have so far developed leukemia; one died of it.
These setbacks were hardly surprising in the early days of gene therapy, White says. Furthermore, the journey from laboratory bench to human use for any drug or biological application is steeply uphill. Promising discoveries can languish as biotech start-up companies may only invest in developing one product at a time. Even when a discovery is fast-tracked, it can take anywhere from one to 10 years or longer to pass through all of the investigative stages and reach the FDA for licensure. Then more time passes while the government reviews it. Ultimately, for new drugs, for example, only one in 100 make it to the marketplace.
The highly publicized clinical tragedies choked initial optimism about gene therapy, increasing caution among scientists and clinicians, and slowing progress. In that less enthusiastic environment, "the problems associated with gene therapy took about 10 years to understand and resolve," White said.
While researchers on both sides of the Atlantic were studying the underlying issues brought to light by the clinical trials, a trio of American women was mobilizing for a personal life-and-death struggle. Amber and Rachel Salzman and their sister Eve Lapin committed to speed up research on behalf of children born with adrenoleukodystropy or ALD, a disease of the fatty insulation (myelin) surrounding nerve cells. ALD is an inherited disorder, invariably fatal in children about two to five years after diagnosis. (ALD was the subject of the 1993 Oscar-nominated film Lorenzo's Oil starring Nick Nolte and Susan Sarandon.)
In December of 2000 Lapin, of Houston, learned that her son Oliver had ALD and probably only a few years to live. After further testing, both Lapin's younger son and Amber's baby boy had the biochemical marker for ALD; both mothers were carriers. Rachel, a veterinarian by training, was not a carrier.
Amber's son was only a year old, yet she understood the situation was urgent. "We knew we had only four or five years to find a cure," she said. Two months after receiving the dire news, Amber and Rachel met in Paris with scientists Patrick Aubourg and Nathalie Cartier of Inserm, a French research institute, to learn about the latest work on ALD and gene therapy in lab animals. And within three months of Oliver Lapin's terrible diagnosis, the three sisters formed the Stop ALD Foundation, supporting scientific collaboration leading to effective treatments and a cure for ALD.
At that time Amber Salzman was a senior executive at GlaxoSmith- Kline. She and Rachel started consulting experts within and outside her company, and "read through every journal." It was Amber's GSK boss, Tachi Yamada, who urged her to explore the use of gene therapy. And in 2001 they began meeting with scientists to do just that.
As she learned more about it, Amber Salzman recognized that a gene therapy solution could not come soon enough to help her son or nephews. Indeed, Oliver died in 2004 at age 12. His younger brother, Elliott, and her son, Spencer Barsh, were candidates for traditional bone marrow stem-cell transplants, the only existing treatment option, but no marrow matches were available. In 2002 the two young cousins received stem cell transplants from umbilical cord blood. (Use of cord blood, which contains early, undifferentiated cells, mitigates the body's immune response to foreign marrow.)
However, such transplants carry their own set of serious risks, most particularly the enduring possibility of an immune system reaction to foreign transplanted cells. Spencer, now 9, "almost died three times and it was a brutal procedure" involving three months in the hospital, Amber says. Today he is healthy and suffers no lasting ill effects. But his older cousin Elliott, now 15, suffers from graft-vs.-host disease and uses a wheelchair.
Having gone through the harrowing traditional transplant process "we felt very strongly that we needed to do something differently," Amber Salzman said. And that something was gene therapy.
Around the time Amber and Rachel Salzman started exploring gene therapy options, Inder Verma was elected the fourth president of the American Society of Gene Therapy (ASGT), thereby becoming a leading spokesman. The sisters were desperate to find a way to address ALD. And Verma had personally developed a viral vector with the ability to enter brain cells – the very types of cells affected by ALD. A meeting was essential.
"The first time I met him, I thought, 'Oh God, I'm talking to this brilliant scientist,' " Amber Salzman said of Verma. "But he puts you at ease. He likes people, likes helping...here was a top-notch scientist with a humanitarian side."
Verma recalled their first meeting, at an ASGT conference in Seattle in 2001. He learned that the French scientists were experimenting with retroviruses to treat ALD in mice. "But that was not very efficient," he said. "I wanted them to use the lentivirus." Amber told him they needed his help.
At their next meeting, shortly after the 9-11 tragedy, convened at the Hilton at a London airport, Aubourg and Cartier were also in attendance. "They showed me the data," he said. "I could see they needed a better and more efficient vector." Convinced the French group needed his lentiviral vector, Verma went back to Cell Genesys and helped persuade the company to make the vector of the quality required for human treatment. (Ultimately Cell Genesys licensed Verma's lentivirus technology to San Diego's Invitrogen to be marketed to other researchers as lab experiment kits; these kits have been widely used. And in March of 2008, Cell Genesys sold the technology to another company.)
Back in 2001, the lentiviral vector had never been used in people. It was expensive to prepare. And Verma was asking Cell Genesys to do all this to treat a rather rare disease. "But it appeared that it was well worth the trial," he said.
Over the past eight years the Salzmans have pushed and prodded to move the science, the scientists and the relevant companies forward. "We were pretty annoying," says Amber now, sounding only slightly rueful. The sisters strongly believed it was their role to make sure the scientists continued to talk to each other, "connecting the people and the pieces." "To Inder's credit, when I called him, he'd call me back within 24 hours," Amber said. "He was so engaged, and so patient at both a scientific and personal level. I feel like he's a member of my family.
"While he was very helpful and supportive," she continued, "he was always concerned about the scientific rigor of the work." Verma demanded, for example, that Aubourg publish his animal data for peer review. "I don't underestimate how much Inder's involvement helped us," she said of Verma.
It took several years for the French scientists to perform the necessary preclinical work with animals and for Cell Genesys to prepare a lentiviral vector suitable for use in a human clinical trial. Finally, in 2006 and 2007, two 7-year-old boys in Spain, both ALD patients with brain lesions characteristic of the disease, received the gene therapy treatment. It was the first time the lentiviral vector was used in humans who did not already have HIV infection.
The beauty of adding gene therapy is that it allowed each boy to be his own stem cell donor, correcting his own cells and eliminating the need to find matching bone marrow with the potentially lethal effects of foreign cells. The treatment began with the removal of some of each patient's bone marrow stem cells. Then the lentiviral vector was used to deliver healthy genes into the cells in a laboratory procedure. The treated marrow cells were then injected back into the patients, so each received his own improved cells.
In a Nov. 6, 2009 paper in Science, Aubourg and Cartier published their initial very promising results, triggering a media barrage. "Gene therapy makes major stride in Lorenzo's Oil disease," declared the Los Angeles Times that same day. "Gene therapy turnaround," said The Scientist on its newsblog. "For Gene Therapy, Seeing Signs of a Resurgence," The New York Times reported. "Gene Therapy for Fatal Brain Disorder 'Just the Beginning'," suggested U.S. News and World Report.
Amber Salzman, who is now the CEO of Cardiokine Inc., a biotech company developing heart medicines, expressed considerable satisfaction with the early results of the gene therapy trial. The young Spanish boys needed only about a month in the hospital and experienced a far gentler procedure than her own son, she said. The gene therapy has arrested the diseaseand both boys in Spain are leading normal lives. (A third boy in Spain received the therapy more recently and is not yet two years post-procedure, but is also doing well.)
Salzman said she is personally persuaded that the lentiviral vector is both safe and effective, converting enough nerve and brain cells to healthy function to make the outlook bright for these children. Her next goal is to expand clinical trials, both in Europe and in the U.S., treating enough patients to definitively show that gene therapy should be "the new standard of care for ALD." Once that happens, insurance companies will pay for the treatments and many more patients can be helped.
Donald B. Kohn, professor of Microbiology, Immunology, Molecular Genetics and Pediatrics at UCLA, warmly greeted the news from France. "Use of the lentiviral vector is a major milestone, advancing the technology," he said. The preliminary results indicate that 15-20 percent of the patients' nerve cells received the healthy genes, and that level of efficient gene transfer "is really unprecedented for a clinical trial."
Kohn, a leader in the gene therapy field, said we don't know why effectively treating far fewer than 100 percent of the nerve cells is sufficient to change the health of a patient, but one can speculate that the number of healthy cells making the proper enzymes is adequate to rescue the patient's other defective cells.
Demand for the lentiviral vector seems destined to increase, as a number of other clinical trials involving other diseases get under way. Kohn was recently awarded a $9 million California Institute for Regenerative Medicine (CIRM) grant to develop a clinical gene therapy trial to treat sickle cell anemia using a lentivirus. He thinks the vector might also be suitable for gene therapy to treat other blood diseases and tissues in the eye.
"The work done by Dr. Verma and his team at the Salk was really pioneering, representing a quantum leap forward," Kohn said. And though it took a decade to go from lab to curing patients, Kohn called that "pretty quick" to address the "learning curve and testing" necessary for human use.
Verma is aware of a number of other clinical trials using the lentiviral vector, including two in London to treat Parkinson's Disease and Retinitis Pigmentosa, an hereditary cause of blindness. In the U.S. the vector is being used to treat SCID (Bubble Boy Disease) and a number of bloodbased diseases including sickle cell and Chronic Granulomatous Disease (CGD), a group of hereditary diseases of the immune system.
Meanwhile, Amber Salzman, Eve Lapin and Rachel Salzman (now a freelance medical analyst who advises companies on diverse therapeutic areas) have an even broader agenda. They want to leverage a newborn screening initiative at the Kennedy Krieger Institute in Baltimore, MD to make it a routine requirement. Babies are not currently tested for ALD at birth and by the time it is diagnosed, it is often too late to help the children, she said. "Doctors won't screen for diseases for which they don't have known safe therapies," Salzman explained, and traditional bone marrow transplants are considered dangerous. She believes the use of gene therapy will change the treatment landscape and make routine testing worthwhile.
And while Salzman, her sisters and the French researchers all deserve major credit for the parts they have played in the ALD treatment story, Amber is quick to bring the focus back to Verma, saying it is "important that he be recognized" for both his tremendous scientific achievement and his interpersonal skills.
"He knows how to bring people together," Salzman said, marveling at Verma's particular ability to remember the names of her children and inquire about them. Not all scientists know how to get basic discoveries to patients, she said, but he can do it.