Gene therapy rescues mice with SMA

Reversing a protein deficiency through gene therapy can correct motor function, restore nerve signals and improve survival in mice that serve as a model for spinal muscular atrophy, new research shows. The research is published online in the journal Nature Biotechnology (february 28, 2010).
The researchers used an altered virus to deliver a portion of DNA that makes the SMN protein into the veins of newborn mice ranging in age from 1 to 10 days old. Newborn mice that received the treatment demonstrated near-normal motor function (movement) and brain-to-muscle signaling, as well as a dramatic increase in length of survival, with most living upwards of 250 days as compared to untreated mice, which had an average life span of 15 days. The earlier the therapy was administered, the better the result. No effect was seen from gene therapy treatment given to SMA-affected mice at 10 days of age.

"We're replacing what we know is lost. And we have shown that when you put the protein in postnatally, it will rescue the genetic defect," said Arthur Burghes, professor of molecular and cellular biochemistry at Ohio State University and a senior co-author of the study. "This technique corrects the mice considerably more than any drug cocktails being studied as a potential treatment in humans."

The scientists used a special form of a virus (AAV9) to deliver the SMN protein to nerve cells in the mice. This virus still has the capability to infect cells but has been altered so it will not copy itself and cause illness in humans, said Brian Kaspar, an investigator in the Research Institute at Nationwide Children's Hospital and assistant professor of pediatrics at Ohio State, also a senior co-author of the study.
Kaspar's lab previously determined that this particular viral vector can cross the blood-brain barrier, a characteristic that is required to ensure this protein reaches nerve cells in the spinal cord. The research group demonstrated that effect by intravenously injecting some of the disease-model mice with a green fluorescent protein that functioned as a marker of where the virus traveled in the body.
Ten days after the injection, 42 percent of spinal motor neurons in these mice showed that they contained the fluorescent protein. Similarly, mice with spinal muscular atrophy that received the SMN protein via the viral vector when they were 1 day old showed increases of the protein in the brain, spinal cord and muscles within 10 days, though the levels remained lower than the levels of SMN in normal mice. Those higher levels of the protein appeared to be sufficient to reverse effects of the disease, Burghes said. That is significant because, based on mouse data, the disease is believed to affect people with SMN levels below about 20 percent of normal. But people with only 50 percent of the expected amount of the protein in their motor neurons do not have the disorder. In addition, a single gene therapy treatment appears to reverse the disease, as opposed to drug treatments under investigation that might elevate SMN protein levels but would require a lifetime of taking medication.
In this study, the researchers tested mice with SMA after the treatment with the protein for their ability to roll themselves upright and for the presence of electrical signals from nerve cells to muscles. Within 13 days after the injection, 90 percent of the treated mice had the muscle coordination needed to right themselves as quickly as normal animals. By this time, untreated SMA mice already were suffering symptoms that left them unable to right themselves. The day-1 treated mice also were nearly identical to normal mice in their ability to run on a wheel. In the case of restored nerve impulses, 90 days after the gene therapy, there was no difference in nerve pulses between the treated SMA mice and normal mice, which indicated that the nerves to muscle developed correctly. The treated SMA mice also gained weight and lived substantially longer than untreated mice with the disease, and some mice were still alive when the paper was submitted 250 days after they received the treatment, Kaspar said. Improvements this dramatic were seen only in the mice that received SMN on their first two days of life. Later delivery reduced the impact of correction.
The results indicate that a "window of opportunity" exists during early life in which intravenous injection of AAV9 containing the SMN gene can have optimum therapeutic benefit. "We don't yet know the exact window of when it is capable of getting into the right cells in a human. Is it a month after birth, or a week after birth? That's still a question," Kaspar said. Complicating this issue is the fact that symptoms of spinal muscular atrophy aren't typically apparent in infants, and the only existing newborn screening technique has not been implemented because it is considered prohibitively expensive. The study results suggest "two critical points," said study team member Arthur Burghes. "First, SMA is not a disorder of early (prenatal) development; therefore, SMN correction can have an effect when delivered after birth in these mice. Secondly, the connection of nerve to muscle is corrected and functions in a normal manner."
The researchers hope to progress to human clinical trials of this gene therapy technique as soon as the requisite toxicology experiments are in place and federal regulators will allow. They hope to treat the first patients within two years.
To help move the treatment toward clinical testing in humans, the investigators tested the ability of the AAV9 viral transporter to reach the spinal cord in a nonhuman primate that did not have a form of SMA. They inserted the gene for green fluorescent protein (GFP) into the AAV9 delivery vehicle and administered it to a macaque monkey via intravenous injection. Analysis after 15 days revealed extensive GFP activity in the spinal cord, indicating successful systemic delivery.
"Ultimately we are trying to determine whether we see the same potential to target motor neurons and other cell types in larger species closer to humans," Kaspar said. "We have great results in a nonhuman primate when the virus was delivered at postnatal day one. We are now looking to determine the time point at which we can target motor neurons most efficiently." "We have assembled a clinical working group and are pursuing these studies to determine safety, dosage and window of opportunity as we plan the first clinical trials," said study team member Kevin Foust. "We are also working to develop an improved virus that is capable of targeting motor neurons in older mice and nonhuman primates in order to extend the potential therapy to older patients." Clinical trials in humans will be necessary before the U.S. Food and Drug Administration (FDA) can consider this technique as a treatment for SMA patients. The positive results from testing in the monkey should help pave the way for such trials, as that animal's genetic similarities to humans indicate an increased likelihood that the delivery method may work in humans.

Nature Biotechnology article

(sources: Ohio State University, MDA, FSMA)