How MDA-supported research to counteract a complement protein and rev up regulatory T cells may improve MG treatment
"I’m on CellCept, prednisone, Mestinon and IVIG every three weeks," says 38-year-old Rachel Pegram. "Prednisone, which I have taken for more than 22 years now, has very rough side effects. It has caused weight gain, diabetes and glaucoma. I cannot say I have ever gone into remission without drugs, but I believe I have been in a drug-induced remission. I have spent a lot of time in hospitals for IVIG treatments for crises."
The Neuromuscular Junction in Myasthenia Gravis
Activating signals are transmitted from nerve cells to muscle cells via acetylcholine (ACh). After ACh leaves a nerve cell, it lands on docking sites called acetylcholine receptors (AChRs), which tell muscle cells to contract. ACh is normally broken down by an enzyme called AChE. Drugs that counteract this enzyme prolong the acetylcholine signal. In most people with myasthenia gravis (MG), antibodies from the immune system destroy AChRs, which become antigens (targets). Drugs that alter the immune system can protect AChRs to some extent. Other neuromuscular junction antigens that can trigger MG are proteins called MuSK and LRP4 (not shown).
The disorder Pegram is describing and that she’s been dealing with since the age of 13 is myasthenia gravis (MG). It belongs to a group of disorders known as "autoimmune" (self-immune) diseases, in which the immune system mistakenly attacks the body's own tissues instead of confining itself to fighting infections. In myasthenia gravis (literally "serious muscle weakness"), the target of the misguided immune system is the neuromuscular junction, the place where nerve fibers meet muscle fibers and activate them through a chemical called acetylcholine.
Most MG patients have fluctuating weakness, which can be life-threatening when it involves the breathing and swallowing muscles, because their immune systems target docking sites for acetylcholine — the acetylcholine receptors.
A minority have similar symptoms because their immune systems attack proteins at the neuromuscular junction known as MuSK and LRP4.
The disease is controlled, albeit imperfectly, by a combination of treatments, including drugs to prolong acetylcholine signaling, such as pyridostigmine (Mestinon); drugs to suppress the immune system, such as prednisone and myocophenolate mofetil (CellCept); a procedure to filter unwanted immune system proteins from the blood (plasmapheresis); and a procedure to infuse new immune system proteins (intravenous immunoglobulins (IVIG, Gammagard and other brand names). Surgical removal of the thymus, an immune system organ located in the chest, is another immune-modifying treatment that is frequently recommended.
"Patients do benefit from chronic suppression of the immune system, but there’s a significant downside," says Premkumar Christadoss, an MDA research grantee at the University of Texas Medical Branch in Galveston, where he's a professor in the Department of Microbiology and Immunology.
For one thing, he says, the immunosuppression makes it difficult for patients to fight infections. And prednisone, a mainstay of MG treatment, comes with a host of unwanted effects — the ones mentioned by Pegram and others, such as osteoporosis, thinning of the skin, high blood pressure and changes in thinking and mood.
That's why Christadoss and others want to improve on current treatments for MG, making them more effective, more specific, more dependable and faster-acting, with fewer unwanted effects.
Christadoss and his colleagues have chosen to work on a part of the immune system known as "complement," a group of small proteins found in the blood that swing into action when the immune system calls on them. Like other parts of the immune system, complement is useful most of the time. But it also can be a problem, helping the immune system to mount a misguided attack on the body’s own tissues in MG and other autoimmune diseases.
In MG, it’s well known that complement plays a role in the immune attack on acetylcholine receptors. (Christadoss says it also may play a role in MG related to an attack on LRP4 but probably not in MG related to MuSK targeting.)
The many proteins of the complement system normally circulate in an inactive state. When activated, they signal each other in a chain reaction that’s often called the "complement cascade." The end result of the cascade is often destruction of cell membranes, such as those in which acetylcholine receptors are embedded. If one component of the complement cascade is missing, the components that follow it can’t be activated.
Christadoss and his colleagues have chosen to focus on suppressing a complement protein called C2, counteracting it with a type of compound called a "small interfering RNA," or "siRNA."
The Immune Response
An immune response starts when an antigen-presenting cell, such as a macrophage or dendritic cell, sees an antigen (such as the acetylcholine receptor in myasthenia gravis) and then engulfs and digests it (1). Antigen-presenting cells then display pieces of the antigen to killer T cells or helper T cells of the immune system (2). Helper T cells can show the antigen to B cells (3), which then produce antibodies (4). These antibodies can directly attack the antigen and can also activate the complement system (5), which also attacks the antigen. Helper T cells also secrete cytokines (6) that stimulate macrophages, dendritic cells and other T cells to augment the immune response and cytokines that stimulate regulatory T cells to dampen the immune response.
C2, says Christadoss, is expressed at a very low level in the complement cascade, making it possible to block it without using much siRNA. It’s also active only in the branch of the cascade that does the most damage in MG — so counteracting it leaves the other branches intact to fight infections.
"Partial suppression of C2 production is enough to control the disease in mice," says Christadoss, referring to mice that mount an immune response to their acetylcholine receptors and develop a disorder that looks a lot like human MG.
In MDA-supported research published 2013, Christadoss and other investigators showed that mice in which the MG-like disease was well underway benefited greatly from treatment with siRNA against C2. Animals treated once a week for five weeks showed significantly improved strength, more functional acetylcholine receptors and diminished attacks on muscle cell membranes.
Perhaps most important for patients, siRNA treatment is target-specific and showed no obvious side effects, at least in the mice. What's more, the mice appeared to have normal immune systems in every respect, with full ability to fend off infections.
In 2011, Christadoss founded Immune Globe, a biomedical startup company whose goal is to develop new products to treat autoimmune diseases, particularly MG. If all goes well, he hopes to have an siRNA anti-C2 compound to test in humans within the next two years.
"I definitely expect changes in MG treatment within the next few years," Christadoss says. "I'm very optimistic about it.”
Revving up regulatory cells
Muthusamy Thiruppathi, an MDA grantee at the University of Illinois, Chicago, is pursuing another strategy for treating MG — improving the function of cells that naturally dampen the immune response in humans and animals. Known as regulatory T cells — T regs for short — these cells usually kick in to balance an immune response that threatens to get out of hand. In patients with MG, however, the T regs aren’t very effective.
In 2012, Thiruppathi and colleagues reported results of an MDA-supported study showing that exposing T regs from MG patients to a compound called GM-CSF markedly improved their suppressive abilities.
In other MDA-supported research, they found that mice with an MG-like disease responded well to treatment with infused T regs that had been previously treated with GM-CSF. Mice treated with these cells showed improvements in strength and reduced numbers of antibodies to their acetylcholine receptors. At the end of the study, mice treated with GM-CSF-exposed T regs had more receptors than mice treated with T regs not exposed to this compound.
In fact, in 2012, a patient with severe MG who wasn't responding to conventional therapy improved after being treated on an experimental basis with GM-CSF, and laboratory tests showed the compound appeared to have dampened the immune response against his acetylcholine receptors.
Although Thiruppathi cautions that conclusions about GM-CSF's effectiveness can't be drawn from a single patient, the results are intriguing and have prompted him to look into the possibility of a clinical trial using this compound.
(Sargramostim, a drug based on GM-CSF, is approved in the U.S. to treat fungal infections and replenish white blood cells following cancer chemotherapy. However, MDA cautions against the use of GM-CSF-based compounds to treat MG until clinical trials confirm its safety and effectiveness in this disorder.)
So far, Thiruppathi has only been working with samples from patients with the acetylcholine receptor type of MG, but he intends to start working with samples from MuSK MG patients as well.
"Current MG treatments are global and have unwanted effects," Thiruppathi says. "A deeper understanding of exactly how the immune system goes awry is emerging now, and it may be possible to change the approach to the treatment of myasthenia gravis from global, nonspecific immune modulation to focused, individualized therapy."
How to learn more
For more on MG and the immune system, be sure to read these past Quest articles:
The Immune Response: A Powerful Ally, a Formidable Foe
As is true with so many aspects of MDA-supported research, studies on how to direct and tame the immune system in one disease, such as myasthenia gravis, often overlap with studies of this complex system in other disorders. For instance, revving up regulatory T cells may become important in treating amyotrophic lateral sclerosis as well as myasthenia gravis; and developing vaccines that allow the immune system to tolerate therapeutic proteins introduced by gene therapy may help patients with Duchenne muscular dystrophy and other disorders, as well as helping patients with autoimmune disorders like MG to tolerate their own proteins.
Stan Appel is a longtime MDA research grantee and director of the MDA/ALS Center at Methodist Neurological Institute in Houston. In his studies of patients with amyotrophic lateral sclerosis (ALS) and mice with an ALS-like disorder, he’s found that the immune system is at first helpful in the disease and later becomes harmful, the balance shifting from immune cells that suppress inflammation in the nervous system to those that stoke it.
Attempts to suppress the immune system in ALS have a long history, Appel says, but not a successful one. "We got this wrong in the beginning," he says. "If you immunosuppress, you take the whole immune system down. The key issue is, you’ve got to have therapies that will upregulate [increase] protective T regs and downregulate [decrease] the attacking T cells. If the ratio of attacking T cells to regulatory T cells stays the same, even if they’re all lowered, you still have a relative ratio of bad guys to good guys, and that doesn’t work."
Improving the numbers and function of regulatory T cells in relation to attacking T cells is something Appel thinks is worth looking at as a potential treatment for ALS, and he hopes a compound called GM-CSF will prove fruitful as a treatment for myasthenia gravis (MG). "I think it’s exciting and absolutely worth trying," he says. "If it works in this population and doesn’t have adverse effects, there could be carry-over to ALS and possibly other diseases."
Even when the immune system is behaving as it should, it can be a barrier to therapies such as organ transplants, cell transplants and gene transfer therapy, because the system considers the new proteins it sees with these therapies as "foreign" and therefore targets them.
Immune responses against newly produced dystrophin protein (made from dystrophin genes) have hampered gene transfer therapy in boys with Duchenne muscular dystrophy (DMD). And immune responses against the viral shells used to deliver therapeutic genes have occurred in other muscle gene transfer experiments, as well as in cell transplantation trials.
Immunologist Lawrence Steinman at Stanford University has MDA support to figure out how to get around these obstacles to new therapies.
He and his colleagues have engineered what they refer to as a "tolerizing" vaccine. Unlike most vaccines, which stimulate an immune response against a specific protein, Steinman’s tolerizing vaccines are designed to cause the immune system to tolerate a specific protein. Proteins targeted by the immune system are called "antigens," and Steinman says the holy grail for treating an unwanted immune response would be “antigen-specific” therapy.
Recently, he and his colleagues devised an antigen-specific, tolerizing vaccine that successfully treated type 1 diabetes, a disorder in which the immune system attacks the insulin-secreting cells of the pancreas.
They made the tolerizing vaccine by inserting DNA sequences into the gene for insulin and packaging the modified gene as an injectable drug. Patients treated with the drug showed evidence of better function of their insulin-producing cells without any unwanted effects.
Steinman, who has recently started a biotech company called Tolerion, wants to broaden this antigen-specific tolerizing approach to other autoimmune disorders, including perhaps MG, and wants to use it to tolerize DMD-affected gene therapy recipients to dystrophin.
While some in the field say what he’s trying to do is too hard, Steinman doesn’t agree. "If people don’t try, they’re never going to get there," he says. With MDA support, he plans to continue trying.