A new study published in the journal Nature Medicine on December 23, 2019, shows how to silence a defective gene responsible for the degeneration of the motor neurons in Lou Gehrig’s disease, or amyotrophic lateral sclerosis (ALS), in adult mice. This could help prevent disease progression if the condition is already symptomatic, and even suppress the illness if it has not yet started.
The current study used a vector, or agent, capable of silencing a particular gene implicated in the motor neuron disease called ALS. The result of this therapy is to either arrest or prevent neuronal degeneration depending on whether the symptoms have already appeared, or it is given prior to disease onset.
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Amyotrophic lateral sclerosis
A rare disease, ALS hits over 5000 Americans each year, and about 30,000 people in the US have this disease at present. ALS is a fatal condition caused by degeneration of the motor neurons in the spinal cord that connect the brain with the muscle cells responsible for voluntary action. Since these nerve cells no longer function, skeletal muscle cells that initiate movement of body parts, speaking, swallowing and breathing, are affected progressively by their lack of nerve supply.
The only thing that can be done at present for these patients is symptomatic treatment. most patients live less than 2-5 years from the date of diagnosis.
ALS can occur in a sporadic or familial form, the former being the most common form by far (90% to 95%). The 5% to 10% that are familial are inherited, due to the occurrence of one or more of over 200 mutations of the SOD1 gene.
This gene is one that encodes the protein superoxide dismutase (SOD), an enzyme that breaks down toxic superoxide radicals that are generated during the cell’s normal metabolism. Earlier work indicates that when the SOD1 gene is mutated, superoxide radicals may not be removed efficiently, or other changes in superoxide metabolism may occur that lead to injury and death of motor neurons. The end result is ALS.
To counter this effect, the scientists introduced shRNA, which is a synthetic molecule engineered to silence the gene target or turn off its expression. This shRNA is delivered to the cell inside a harmless virus called AAV (adeno-associated virus).
Earlier researchers had tried the effect of introducing the shRNA-carrying vector into the blood directly, or into the cerebrospinal fluid (CSF) that bathes the central nervous system. They found that the injections were capable of delaying the progress of the disease but not of stopping it altogether. In fact, the mice treated in this way died quite soon after this treatment was given.
In the current study, the AAV vector containing the shRNA was injected at two places in the spinal cord of the adult experimental mice in which the ALS-associated gene defect was present. One group of mice did not yet have symptoms while the other group of animals had already begun to show signs of illness. The injection was given in the subpial space. The pia mater is the delicate film-like membrane that covers the surface of the brain, dipping into all its folds like a thin transparent plastic sheath.
The results were amazing. A single injection into the subpial space reduced the level of nerve cell degeneration in the mice carrying the gene defect, before the onset of symptoms, causing them to appear and test normal with respect to neurological function. In fact, it appears that the injection resulted in almost 100% protection of the motor neuron cells and other related parts, such as the neuromuscular junctions.
If the mice already had symptoms of neuron degeneration, the injection prevented further damage to the motor neuron cells and halted the progress of the condition. In both cases, therefore, the mice showed no adverse effects throughout the study period.
Even at one year following treatment, the mice remain free of any discernible side effects.
The scientists repeated the same injection technique with the same vector in adult pigs, because their spinal cord size is remarkable like that of humans. They wanted to learn if the approach worked safely and effectively in these large mammals. The device they used was one adapted for human use. They found that indeed the technique was both reliable and uncomplicated.
Implications and future directions
Researcher Martin Marsala says, “At present, this therapeutic approach provides the most potent therapy ever demonstrated in mouse models of mutated SOD1 gene-linked ALS.”
The researchers will now try to validate the effectiveness and safety of the treatment using large experimental animals, and to establish the dose required for its optimal and safe effect. They hope to confirm the safety of the technique in large animal models that are more like humans in their physiology, since this is essential to conducting a clinical trial of this approach in the future.
The scientists say that the fact that the new treatment can be given by subpial injection of the silencing shRNA within the AAV9 vector into the spinal cord makes it an attractive option for other hereditary types of ALS and other diseases due to spinal degeneration, like C9orf72 mutation-linked ALS, or in certain lysosomal storage diseases. This is because the therapeutic agent, whether a corrective gene or a gene-silencing cellular apparatus, can be given straight into the parenchyma of the spinal cord.
Bravo-Hernandez, M., et al. (2019) Spinal subpial delivery of AAV9 enables widespread gene silencing and blocks motoneuron degeneration in ALS. Nature Medicine. doi.org/10.1038/s41591-019-0674-1.