Amyotrophic Lateral Sclerosis Research Program 2012 Investigator Vignette

AAV-Mediated Inclusion Formation as a Novel Gene Therapy Strategy for ALS

Investigator: Ole Isacson, MD, PhD; McLean Hospital

ALS is a progressive neurodegenerative disease, and when a patient has the signs of ALS or Lou Gehrig's disease as it sometimes is called, up to 50% of those so-called motor neurons have died. So it's already quite late in the disease process.

So, the research goal for my DoD grant is to try to find ways of treating disease very early or even preventing it.

The most linear way of thinking about neurodegenerative diseases or all disease for that matter is to look for one thing that causes the disease. For example, in ALS there is a gene called SOD, superoxide dismutase that has a genetic defect. But what usually happens is that single mechanism is not the only reason a cell dies, because that gene is expressed in every cell in the body. But ALS or neurological diseases are not skin diseases or liver diseases for that matter.

So we took a different approach, which was to look at how each cell deals with the mutation of the protein and tried to discover why certain cell types are affected by disease.

And the way that developed was, we had a rat model of ALS called an SOD-mutant rat, which is the same gene that mutates in familial ALS and when the rat started to show symptoms of disease we sacrificed them and studied their nervous system. And just like in ALS, we found that 50% of the nerve cell had died in the spinal cord, but we also found that the motor neurons that control the eye muscles or also the facial muscles were relatively spared or protected.

We then used a laser capture technology in which you shine a laser on each one of these nerve cells, maybe 1,000 are contained in each of these regions-and you can then collect their characteristic expression for each cell. And that meant that we could study each motor neuron that's affected in ALS and actually dividing them up in populations, and so we could isolate the nerve cells that were most vulnerable and the ones that were most resistant and then compare their genomic profile, we call them genetic profiles.

What you see here in the red and blue are each little dots there are different genes and their level is high or low depending on their color. So in the protected regions like your eye muscle control you see there's an up-regulation in red or compared to others that are more vulnerable in blue.

And so by analyzing these patterns of genes, we came up with genes that seemed to correspond to very healthy or strong neurons even in the face of this disease process. And one of the genes that stood out-we knew was a trophic factor, an insulin growth factor two- was overexpressed, so about two- to threefold in neurons that were protected versus those that were more vulnerable.

So we took this protein actually and poured it onto these motor neurons you see here in-in red and green. And indeed when we stressed the neurons with a toxin that simulates ALS all of the neurons were protected in the dish versus about a 50% loss, which is what you see in the disease as well,

We then developed a plan for how to induce or make more of an expression of such proteins in these neurons. And the DoD program that was funded basically allowed us to take already approved drugs, they're called FDA-approved drugs, they're called a library sometimes, and test them on nerve cells and see which one of these already nontoxic drugs could induce these protective proteins.

One of the most important parts of our research program on ALS is to do a high-throughput screening. And what we did was to build a cell that had a fluorescent color marker that showed us when the IGF-II neuroprotective protein was turned on.

And that we did in collaboration with the lab of drug development which is a Harvard-based laboratory that helps with drug discovery. So in that work we're now studying new chemical compounds that could drive this protective protein in cells and some of which could become new drugs in the future.

At the same time though we're using already established drugs which brings down the discovery time by about 90% and the cost also by 80 to 90% because most drugs fail in clinical development because they're toxic, not to the cells that you try to treat, but usually to other cells and organ systems in the body.

So our program now has identified a molecule, a drug already on the market for various other indications that drive the expression of this IGF-II. And we're now going to test that drug on rat models of ALS and see whether it can prevent or delay the disease in those animal models. So this new idea, which is called repurposing of drugs, is something that we really have followed in this DoD program and we think it's very promising.