New Hope For ALS Seen With Genetic Techniques, Growth Factors

Research suggests that implanting cells to support the motor neurons that die off in ALS can help stave off the disease. Another group has developed technology to “silence” the mutant gene that causes ALS. And a third group of scientists found that a support factor rescues motor neurons and could be of value in treating ALS.

Also called Lou Gehrig's disease, ALS is a progressive neurological disease that paralyzes its victims by harming nerve cells that control muscles. The estimated 5,000 Americans newly diagnosed each year experience progressive muscle weakness that can hinder movement, speech, and even swallowing and breathing. Most cases of the disease are “sporadic”—that is, they occur out of the blue. About 5 percent of cases are inherited, and result from mutations in genes expressed in all tissues of the body, but that mysteriously affect only neurons that control muscles. The inherited form is clinically identical to the sporadic form.

Don Cleveland, PhD, and his colleagues at the University of California at San Diego created a mouse model of ALS with a mutation in the enzyme copper-zinc-superoxide dismutase (SOD1), which causes about 20 percent of the inherited forms of ALS, and about 1 to 2 percent of total cases.

They manipulated the mice's genes to give some cells mutant SOD1 and some normal SOD1. Curiously, the toxic effects of the SOD1 gene mutation have nothing to do with the enzyme's normal function. Instead, the mutation sends motor neurons down a cellular pathway to death by damaging not only the motor neurons but also their surrounding support cells. These support cells, called glia, provide important growth factors to neurons, and keep the surrounding environment clean and safe. When glia cells don't perform up to snuff, the neurons suffer.

The researchers found that when neurons with the SOD1 mutation were surrounded by glia with normal genes, the glia were able to stave off the disease. But when mutant glia cells encircled genetically normal motor neurons, the motor neurons were attacked by the glia.

The finding could someday lead to a new therapy for treating ALS. A major line of attack against ALS is to replace motor neurons with stem cells, which would have to re-grow long distances to the muscles in order to work. “In place of this herculean task, we can supply normal supporting cells to prevent the degeneration of motor neurons,” says Cleveland.

The researchers will next investigate the mechanism of the glia cell attack. The team found clues that the glia no longer keep the environment safe for neurons, and that they may mount a lethal inflammatory response against the neurons.

In other work, scientists at the Swiss Federal Institute of Technology of Lausanne (EPFL) tackled a different genetic approach against the familial form of ALS that targets the SOD1 gene. Patrick Aebischer, MD, and Cédric Raoul, PhD, used a technology called RNA interference to shut off the toxic SOD1 mutant gene that causes the motor neuron disorder in ALS mice models.

RNA, or ribonucleic acid, is a molecular intermediate between DNA and protein. Each DNA gene holds the code for production of a particular protein. That code is read, or transcribed, by cellular machinery to make a copy of messenger RNA (mRNA). Each piece of mRNA is then translated into the protein. RNA interference is a cellular process that leads to the destruction of a specific mRNA target by naturally occurring RNA pieces.

To inhibit SOD1 mutant gene expression, the researchers engineered a virus to deliver molecular instructions that activate RNA interference. Once inside the cell nucleus, the virus sends a genetic signal to destroy mutant SOD1 messenger RNA. By destroying the RNA, the mutant gene is prevented from becoming a protein and carrying out its devastating effects.

By injecting the virus directly into the spinal cord, Aebischer and Raoul aimed their weapon at the affected area. This is among the first studies in which RNA interference has been used as a therapy against a neurodegenerative disease, Aebischer says.

“We targeted the cause of the disease by knocking down mutant SOD1 in the spinal cord,” Raoul says. “This helped delay the first signs of motor impairment and slowed disease progression.”

Because the normal copy of the SOD1 gene performs important duties in cells, the researchers built a normal copy of the gene into the virus they delivered to the cells. This feature—removing a harmful version of a gene while replacing it with a normal one—may allow a similar technique to be used in many other diseases affected by other genes.

“This work opens a therapeutic avenue to neurodegenerative disorders including Parkinson's or Huntington's disease,” says Aebischer.

In another study, Erik Storkebaum and Diether Lambrechts, PhD—working in the laboratory of Peter Carmeliet, MD, PhD, at the Catholic University of Leuven in Belgium—rescued the motor neurons attacked by ALS by providing them with a key support agent called vascular endothelial growth factor (VEGF).

VEGF was first discovered as an agent that helps grow blood vessels. It has more recently been shown to support neurons. In both rodents and people, a low level of VEGF may be a risk factor for ALS . So Storkebaum and his colleagues tested whether raising VEGF levels could better protect against ALS in a mouse model.

The investigators first delivered VEGF to the brains of ALS rats with a surgically implanted needle. The compound was added to the cerebrospinal fluid, which constantly “washes” the entire brain and spinal cord.

“When VEGF was administered one month before the disease symptoms showed up,” Storkebaum says, “treatment delayed the onset of paralysis, improved motor performance, and prolonged life span by about 15 percent.”

In a second experiment, the researchers wanted to more closely mimic the human clinical situation, so they waited for motor abnormalities to arise in the rats. At this stage many motor neurons were already lost, yet the VEGF treatment still provided benefits, prolonging the rats' lives slightly.

The researchers then tested whether more receptors for VEGF might give the neurons an advantage. They genetically engineered mice that had ALS and expressed the VEGF receptor in greater numbers than normal. “Overexpression of the VEGF receptor indeed protected against motor neuron degeneration,” says Storkebaum. This result indicated that VEGF could act directly on motor neurons.

Next, they attempted to increase the supply of VEGF to neurons. Motor neurons, which are stricken by ALS, make physical contacts with muscle fiber cells. There, they release chemical messengers that tell the muscles to contract. This point of contact is called the neuromuscular junction.

They used a virus to deliver the gene for VEGF to muscle cells, resulting in increased expression of VEGF protein by the muscle cells. Once the virus was absorbed, the muscle cells started to produce VEGF, making it available to the neurons at the neuromuscular junctions. Once again, VEGF improved the disease course—the onset was later and the mice survived longer. The researchers also found more neuromuscular junctions, evidence that VEGF helps to preserve these crucial points of contact.

In future studies, the team hopes to determine whether the virus containing the VEGF gene can also be taken up by the nerve endings. If so, the neurons might transport the gene back to the cell body and produce VEGF in the spinal cord, where it could provide help to other neurons. Alternatively, the VEGF protein itself might be taken up by the neurons and transported to the cell bodies.

“These results raise hope for a potential new treatment for ALS patients,” says Storkebaum. However, further studies in animals and clinical trials in patients will be needed.