Results of the research, which was carried out at the Ecole Polytechnique Fédérale de Lausanne (EPFL) in Switzerland, will be published in the April issue of Nature Medicine and currently are available in the journal's advanced online publication.

Mutant Silenced, Normal Gene Delivered

In addition to silencing the mutated gene that causes ALS, the EPFL researchers were able to simultaneously deliver a normal version of the gene to motor neuron cells using a single delivery mechanism.

"This is the first proof of principle in the human form of a disease of the nervous system in which you can silence the gene and at the same time produce another normal form of the protein," notes Patrick Aebischer, EPFL President and a co-author of the study.

ALS is a progressive neurological disease that attacks the motor neurons controlling muscles. Although its victims retain all their mental faculties, they experience gradual paralysis and eventually lose all motor function, becoming unable to speak, swallow or breathe.

This harrowing disease acquired the name "Lou Gehrig's disease" from the baseball player who succumbed to it. It has no cure, and its pathogenesis is not very well understood.

Using 'Molecular Scissors'

An estimated 5,000 Americans are diagnosed with ALS every year, and most of these cases are "sporadic," with no identifiable cause. About 5-10% of ALS cases are inherited. Of these, 20% have been linked to any of more than 100 mutations in the gene that expresses the superoxide dismutase enzyme (SOD1).

SOD1 mutations are "toxic gain-of-function mutations," meaning that the protein expressed by the mutated gene has -- in addition to all its normal cellular functions -- some additional function that makes it toxic to the cell.

"Any mutation to the SOD1 gene is fatal to motor neuron cells," Aebischer notes. Recent research also indicates that mutant SOD1 gene expression in neighboring glial cells is also implicated in motor neuron death.

Lead author Cedric Raoul and colleagues targeted the cause of the disease by using RNA interference to silence the defective gene, preventing it from expressing the SOD1 protein.

RNA interference is part of a complex cellular housekeeping process that protects cells from invading viruses or other genetic threats. It works by interrupting messenger RNA as it transfers the genetic code for a protein from the nucleus to the site in the cell where the protein is synthesized.

To trigger RNA interference and silence a gene, short bits of double-stranded RNA are introduced in the cell, where they bind with matching sections of messenger RNA. The cell identifies the resulting messenger RNA strand as faulty, and chops it up. As a result, the genetic blueprint isn't delivered, and the protein never gets made.

"Gene silencing is an example of using "molecular scissors" at its most advanced level," Raoul explains.

Improved Neuromuscular Function in Mice

Raoul and colleagues used RNA interference to reduce levels of mutant SOD1 protein in the spinal cords of transgenic ALS mice. Short strands of RNA that targeted multiple mutated and normal forms of the human SOD1 gene were delivered in a specially engineered lentivirus.

Expression of the SOD1 protein was knocked down in the affected motor neurons and neighboring glial cells, and both the onset and the rate of progression of the disease in the treated mice were substantially reduced. In addition, the mice showed a significant improvement in neuromuscular function.

"This is the first demonstration of therapeutic efficacy in vivo of RNA interference-mediated gene silencing in an ALS model," notes Raoul.

Because the normal form of the SOD1 protein may be necessary for the survival or function of adult human motor neurons, the Swiss researchers designed a gene-replacement technology that allows the knock-down of all mutant SOD1 forms while permitting the expression of a normal type SOD1 protein that is resistant to RNA interference-based silencing. Both these effects are expressed long-term via delivery by a single lentiviral vector.

Hope for Parkinson's, Other Nerve Diseases

Aebischer is optimistic about the future of gene silencing as a potential therapy, particularly in incurable progressive neurological diseases such as ALS.

"I would not be surprised to see, in the next ten years, this technology used for treating diseases of the nervous system, particularly diseases that involve toxic gain-of-function, such as inherited forms of Parkinson's disease or Huntington's disease," notes Aebischer.

"But it's important to note that the safety of delivering lentiviral vectors to the nervous system will have to be carefully examined prior to treating patients," he says.