Gene therapy for arthritis and other non-terminal, debilitating conditions and diseases is both feasible and safe, report researchers who conducted the world's first such test on the approach in patients with advanced rheumatoid arthritis.
The results, published in this week's online edition of the Proceedings of the National Academy of Sciences (PNAS), indicate that introducing a new gene has the potential to block the destructive inflammation process that takes place within arthritic joints. At a time when setbacks have cast doubt on the future of gene therapy, the results also provide assurance that it can be performed without causing undue harm to patients.
The clinical trial, which was conducted at the University of Pittsburgh School of Medicine between 1996 and 1999, involved nine women who had genetically modified cells injected into their arthritic knuckles and marked the first time a gene was introduced into a human joint. The researchers say their results showing that successful gene transfer can target joint inflammation open the door to the development of improved gene-based therapies for both rheumatoid and the more common osteoarthritis, which together affect about 66 million people in the United States. While the Pitt study used the same vector – an inactivated virus that shuttles a gene into cells – as a more recent French trial for X-linked severe combined immunodeficiency (X-SCID) in which three children later developed leukemia, the researchers point out that this is the only similarity between the two trials. The studies were concerned with different genes and with different targets, and in the arthritis trial, the transduced cells were removed after one week during routine joint replacement surgery. Nonetheless, with safety a key concern, the authors report no clinical side effects up to five years after the procedure and no evidence that the vector, a replication-defective retrovirus called Maloney Murine Leukemia Virus, has since become capable of replication in their patients.
"At the outset, our primary objective was proving the safety of gene therapy, especially for a clinical application to a nonlethal disease, such as arthritis. Once we learned of the adverse events in the X-SCID trial, we decided it was best to extend follow-up of our patients to five years so that there'd be little question about long-term safety," explained Christopher H. Evans, Ph.D., D.Sc., the study's lead author, now director of the Center for Molecular Orthopaedics at Brigham and Women's Hospital and Robert Lovett Professor of Orthopaedic Surgery at Harvard Medical School.
In rheumatoid arthritis, immune system cells called macrophages and lymphocytes colonize the lining of joints where they release proteins called cytokines that modulate communication between the immune cells and synovial cells, the cells that line the joint. As the researchers discovered in earlier animal studies, synovial cells have a receptor on their surface that's a perfect fit for a particular cytokine, interleukin-1, or IL-1. Like a switch, when IL-1 binds to this receptor, the cell unleashes additional biochemical agents, which in turn cause more local inflammation. As inflammation builds, patients experience progressively worse pain and stiffness in their affected joints.
To block this process, the team sought a way to prevent the main actor, IL-1, from binding to the synovial cells, and hence, setting off the destructive chain reaction that follows. In essence, they needed a way to plug up the receptor and found such a device in a gene encoding the IL-1 receptor antagonist, or IL-1 Ra.
Because it was the first trial of its kind and safety was a key concern, the protocol was designed so that the gene would be delivered to cells that had previously been removed from patients and then, after gene transfer, reintroduced several weeks later after intensive screening and determination of gene expression.
"Given that gene therapy was so new and we were dealing with an otherwise healthy patient population, the ex vivo protocol was the most appropriate approach to take at the time," stated Paul Robbins, Ph.D., professor of molecular genetics and biochemistry and orthopaedic surgery at the University of Pittsburgh School of Medicine, who was the study's co-principal investigator and then director of the University of Pittsburgh's Viral Vector Core Facility.
According to their results, joints treated with the genetically modified cells exhibited high levels of IL-1 Ra, indicating successful gene transfer. Clusters of cells that expressed large amounts of the gene were present at the surface of the synovial tissue and produced significantly less of the inflammation-provoking IL-6 and PGE 2 than cells within joints that were not treated with the gene.
The nine women enrolled in the trial were between 49 and 73 years of age and had been living with rheumatoid arthritis for between 10 and 26 years. Each was scheduled for joint replacement surgery involving the four knuckles on one of their hands and one additional joint. In the weeks prior to joint replacement surgery, synovial tissue was removed from the additional joint that required surgery. Cells from the tissue were then cultured for several weeks, after which time half the cells had the IL-1 Ra gene inserted and half remained untreated. Six to seven weeks later, following extensive laboratory screening of the genetically modified cells, patients returned to the clinic, where the cells were injected into four knuckles in a double-blind fashion, with two knuckles receiving the gene-modified cells, and two knuckles receiving injections of cells with no added gene. One week later, at the time of the patient's previously scheduled surgery, the four knuckles were removed for study and replaced with artificial joints.
In their animal studies that preceded the clinical trial, the researchers had noted that a small percentage of injected cells could escape to other tissues within one week. But with five years follow-up, the authors have seen no evidence that these genetically modified cells have caused any adverse events in their nine patients.
Although the ex vivo approach did prove safe and was able to confer gene expression within arthritic joints, the authors contend that it also was tedious, time-consuming and expensive. At this point they would advocate a more direct in vivo approach – introducing the gene directly to affected tissue – using as the vector an adeno-associated virus (AAV) instead of the vector used in the trial reported in PNAS. AAV appears to have a good safety profile and, in animals, facilitates more extended gene expression. The investigators hope to conduct additional clinical trials in both osteoarthritis and rheumatoid arthritis using AAV to transport the gene.
In addition to Drs. Evans and Robbins, other authors are James H. Herndon, M.D., also now at Harvard; Steven C. Ghivizzani, Ph.D., now at the University of Florida; Mary Chester Wasko, M.D., Molly Vogt, Ph.D., Theresa L. Whiteside, Ph.D., Elaine Elder, Ph.D., and Simon C. Watkins, Ph.D., all of the University of Pittsburgh; Matthew M. Tomaino, M.D., now at University of Rochester Medical School; and Richard Kang, M.D., and Thomas A. Muzzonigro, M.D., who are both now in private practice.