Imaging ALS. Neurologist Martin Turner MA PhD FRCP is developing MRI methods to diagnose and monitor ALS. Image: Courtesy of Martin Turner MA PhD FRCP.

ALS is typically diagnosed within 12 to 14 months. But people with ALS do not have that kind of time to waste. Riluzole extends survival only 3 to 6 months. And, a growing number of clinical trials restrict participation to 24 months after the first symptoms appear - leaving people with ALS few opportunities to access potentially beneficial medicines to treat their disease.

Some neurologists suspect, however, that this diagnostic delay might be cut in half by scanning the brain by MRI and looking for tell-tale signs of ALS. Combined structural and functional MRI methods according to recent studies can help distinguish ALS patients – at least from healthy people. And, MRS-based measures of certain neuronal metabolites might help identify people with inherited forms of ALS before they exhibit the first signs of the disease.

What’s more, these techniques might help clinicians to more accurately diagnose people with ALS enabling better management of their disease. Certain structural MRI-based methods might help distinguish people with C9ORF72-linked ALS. Functional MRI measures might indicate certain cognitive challenges. And, certain structural MRI measures might help identify people with a rapidly progressing form of the disease.

ALS Today's Michelle Pflumm PhD talked to University of Oxford neurologist and neuroimaging specialist Martin Turner MA PhD FRCP at the 2012 American Neurological Association meeting to learn more about MRI and its potential for people with ALS going forward.

Can MRI be implemented today to help diagnose ALS?

We have got lots of ways to separate patients from healthy people.  But that’s not the question we’re asking in the clinic. We can see they’re not healthy.  What we want to know is whether they’ve got ALS or perhaps something else.  That’s the real question.

What needs to happen to introduce these methods into the clinic?

We have the candidates.  We now need to translate them by taking people early in their workup - perhaps when they are first referred to electromyography (EMG).  Some of them will have ALS and some of them won’t.  If we scan people then, we can get a sense of how much it adds.  EMG is probably our biggest competitor in terms of making a diagnosis. That’s our benchmark.

No study to date has successfully used these methods to monitor people with ALS over the course of the disease.  Why is it so challenging?

It is what I call the inconvenient truth of ALS.  When we are studying affected patients, what we are effectively seeing is an iceberg rising up out of the water. But what lies underneath, I suspect, is many years of accumulating pathology. It’s been building up for a long long time.   I don’t think we are going to see a great change.  It is hard to detect changes in an already damaged system.

You recently kickstarted studies with University of Miami’s Michael Benatar MBChB DPhil to look for changes in people at high risk for developing inherited forms of ALS.  Why?

I think they’re the group that we need to focus on now.  There might be signatures that are much more important in terms of capturing the essence of the disease.

I think it will be possible to identify changes [in them].  In Alzheimer’s disease, we can pick up all sorts of changes in the brain 20 to 30 yrs before the disease. The bigger challenge is translating those changes to the 90% without clear genetic risk.

Chemical imbalance? Researchers hope to zero in on chemical changes in certain parts of the brain using magnetic resonance spectroscopy to identify people likely to develop inherited forms of ALS and monitor their disease. Image: Govind et al. (2012), PLOS One.

Do you think these studies will nonetheless help people with the sporadic form of ALS (no family history of the disease)?

I think there is definitely great potential benefit. The biomarker signature that we find in a gene carrier without symptoms maybe what we can focus in on and look at in a patient in an early stage. This would allow potential drugs to be started sooner.

Your most recent results, presented here at ANA 12, suggest that a routine type of MRI scan called FLAIR might also be helpful in identifying people with ALS – potentially enable more routine diagnosis of the disease.  Can you tell us about that?

We find that the FLAIR signal is certainly higher in the corticospinal tract of ALS patients and even higher in PLS patients. [The technique] might be helpful in providing objective evidence of upper motor neuron involvement, a cornerstone of ALS diagnosis.

People with PLS often have to wait 4 years before being formally diagnosed.  Do you think that this technique can help speed up this process too?

Absolutely, with the caveat that it is not common.  PLS is rare.  Its only 3% of all cases. But it is important to study because it may help us understand how to slow ALS down.

What needs to happen to put these methods into practice on clinical MRI scanners?

The results are hinting that there is a lot of information encoded in a very routine scan which you can unlock using mathematical tools; software that’s been developed for non-routine scans such as diffusion tensor imaging.  The next step is to take images from a clinical MRI scanner and see whether they give the same information.

What do you think needs to happen to encourage neurologists to adopt MRI methods in general ALS practice?

What we have to show is that they have unique value – that we can get someone in a trial sooner.  I think that is pretty likely that they can deliver on that.

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To learn more about how neurologists hope to use MRI to diagnose and monitor people with ALS, read MRI, Make That A Double. To find out how neurologists hope to use MRS to identify people with inherited forms of the disease more quickly, check out NAA MaRkS The Spot. 

References

Menke, R.A., Abraham, I., Thiel, C.S., Fillippini, N., Knight, S., Talbot, K. and Turner, M.R. (2011) Fractional anisotropy in the posterior imb of the internal capsule and prognosis in amyotrophic lateral sclerosis. Archives of Neurology doi:10.1001/archneurol.2012.1122. Abstract | Full Text (Subscription Required)

Douaud, G., Filippini, N., Knight, S., Talbot, K., and Turner, M.R. (2011) Integration of structural and functional magnetic resonance imaging in amyotrophic lateral sclerosis. Brain 134, 3470-3479. Abstract | Full Text (Subscription Required)

Carew J.D., Nair G., Andersen P.M, Wuu J., Gronka S., Hu X., and Benatar, M (2011). Presymptomatic spinal cord neurometabolic findings in SOD1-positive people at risk for familial ALS. Neurology 77(14), 1370-1375.  Abstract | Full Text (Subscription Required)

Filippini, N., Douaud, G., Mackay, C.E., Knight, S., Talbot, K., and Turner, M.R. (2011). Corpus callosum involvement is a consistent feature of amyotrophic lateral sclerosis. Neurology, 75(18), 1645-1652.  Abstract | Full Text (Subscription Required)

Further Reading

Benatar, M. and Wuu, J. (2012). Presymptomatic studies of ALS: Rationale, challenges and approach. Neurology Abstract |  Full Text (Subscription Required)

Bowser, R., Turner, M.R., and Shefner J. (2011). Biomarkers in amyotrophic lateral sclerosis: opportunities and limitations. Nature Reviews Neurology, doi: 10.1038/nrneurol.2011.151 Abstract | Full Text (Subscription Required)

Turner, M.R. (2011). Towards a neuroimaging biomarker for ALS. Lancet Neurology. 10(5), 400-403. Full Text (Subscription Required)

Patient Resources

The Oxford Study for Biomarkers in MND/ALS (BioMOx).  Contact | Website

The Pre-Familial Amyotrophic Lateral Sclerosis (Pre-fALS) Study. Contact | Website

With the discovery of nearly 20 genes linked to ALS, researchers are unraveling this complex disease at increasing speed.  But, according to recent research, there is much more than a handful of genetic errors in people with ALS that contributes to the destruction of the motor nerves.

The epigenome – the chemical tags that mark which genes are turned on and off in our tissues – is emerging as a new player in neurodegenerative diseases including ALS.  Inherently flexible, the epigenome rapidly evolves to help people adapt to a crash diet, stress or simply a change in scene. But in people with ALS, studies suggest, these changes might actually fuel the progression of the disease.

Emerging drugs that restore or remove these so-called epigenetic marks may have the potential to treat ALS.  But figuring out how to safely reset these chemical switches in motor neurons using these medicines, according to experts, may be tricky.

AcE AcE Baby 

Critical genes are switched on or off in specific tissues in part by adjusting their accessibility to the intracellular decoding machines. Known as acetylation, chromatin packing proteins called histones get pushed aside, allowing these genes to be transcribed and proteins, produced. But in people with a growing number of neurological conditions including ALS, the levels of histone acetylation drops, contributing to neurodegeneration.

 

HDACs Crash.  Histone deacetylases (green) remove acetyl groups (red) rendering the genes unreadable by the transcriptional machinery (dark blue). Image: National Cancer Institute.

Epigenetics first grabbed neuroscientists’ attention in 2001 when University of California Irvine scientists Leslie Thompson PhD and Larry Marsh PhD reported that the loss of histone acetylation in fruit flies contributed to Huntington’s disease. The team found that the fruit flies did not produce a critical histone acetyltransferase (HAT). But treatment with histone deacetylase enzymes (HDACs) blockers to boost acetylation levels stopped the disease in its tracks.

“These results really created enormous awareness,” explained New York Burke Rehabilitation Center neurologist  Raj Ratan  MD, “that small molecule [HDAC] inhibitors that had been developed for cancer might actually have benefit for neurodegenerative diseases.”

Subsequent preclinical studies found that several compounds that broadly block HDACs appeared to slow neurodegeneration.  But researchers still remain unsure why exactly these drugs might be therapeutically beneficial. 

Scientists speculate that HDAC inhibitors may switch on the production of substances that helps nerves withstand stresses associated with these diseases.  A multi-institutional team led by Raj Ratan, for example, found in 2003 that the HDAC inhibitor trichostatin A increased the levels of acetylated transcription factor Sp1 in neurons, boosting the production of proteins that help protect them from the flood of damaging reactive oxygen species (ROS). And in 2006 and 2008, researchers from the National Institutes of Health in Washington D.C. found that HDAC inhibitors sodium phenylbutyrate, trichostatin A and valproic acid encouraged astrocytes to produce so-called neurotrophins that protected cultured neurons from destruction.

Lost In Translation

 

Trichostatin A.  Researchers from University of Southern California reported in 2011 that trichostatin A could reduce motor neuron loss and extend survival of mice experiencing symptoms similar to ALS. 

Researchers first began to suspect that epigenetic changes contributed to ALS in 2003 when scientists reported that levels of a critical histone deacetlyase in mice dropped 70% primarily in degenerating motor nerves.  Subsequent preclinical studies indicated that three broad spectrum histone deacetylase inhibitors – sodium phenylbutyrate, trichostatin A, and valproic acid  – might be useful in treating ALS because treatment delayed clinical onset and/or extended survival of mouse models.  But independent research teams at ALS TDI in Massachusetts were unable to reproduce these findings for any of these medicines.  And, a phase III clinical trial of 160 ALS patients in the Netherlands found that valproic acid at least at a dose typically prescribed for epilepsy, did not appear beneficial.

A critical limitation of today’s HDAC blockers according to some experts is these drugs may simply need to be administered at too high a dose to be effective.

An estimated 3-5% of genes in people are switched on by today’s histone deacetylase inhibitors such as trichostatin A according to experts.  Many of these genes protect the motor nerves from further deterioration.  But many others likely promote their destruction.

Researchers are now working hard to develop potential medicines that block specific “classes” of histone deacetylases in hopes to identify a drug that more specifically switches on neuroprotective agents to improve the safety and tolerability of these medicines.  These include Repligen’s Class I HDAC1, HDAC2, HDAC3 and HDAC8 inhibitor RG2833 which is being developed for the treatment of Friedrich’s ataxia and GlaxoSmithKline spinoff Tempero Pharmaceuticals’ Class IIa HDAC4, HDAC5, HDAC7 and HDAC9 inhibitors, for the treatment of chronic inflammatory diseases including potentially multiple sclerosis. 

Meanwhile, researchers elsewhere are going for the gold hoping to create potential medicines that selectively block specific HDAC enzymes to treat neurological diseases.  MIT neuroscientist Li-Hui Tsai PhD, for example, hopes to create potential medicines for Alzheimer’s disease by generating compounds that specifically block HDAC2 to treat memory loss.

”I think you are really seeing the HDAC inhibitor [field] blossom,” says Ratan.

The jury is still out however whether any of these more selective drugs being developed are therapeutically beneficial in ALS. 

Despicable ME 

 

All about ME.  Researchers discovered that the DNA methyltransferase Dnmt3A may trigger the deterioration of the motor nerves. Image: Structural Genomics Consortium, University of Toronto.

Johns Hopkins neuroscientist Lee Martin PhD however, suspected that there was much more than changes in histone acetylation that occur in people with ALS.  Critical genes could also be inactivated by methylation in motor neurons by DNA methyltransferases, triggering the destruction of the motor nerves. Enzymes therefore, that could also be targeted to treat the disease.

“Seeing that there was promise in histone modifications,” explained Martin, “maybe there could also be some promise in [targeting] DNA methylation.”

To determine whether increases in methylation could also lead to ALS, Martin’s team in the late 2000s measured the amounts of DNA methyltransferases in the brain and spinal cord of patients and healthy people. The researchers found  that the levels of the two of these enzymes, Dnmt1 and Dnmt3A were significantly higher in the CNS of people with ALS, suggesting that hypermethylation of certain genes could indeed be contributing to the disease.

“There was no data to say that DNA demethylation,” however, explained Martin, “could actually cause neurodegeneration.”

So, the Johns Hopkins researchers turned to model systems. In culture, the researchers found that increased levels of Dnmt3A triggered the degeneration of mouse spinal cord motor neurons. And in mice, treatment with the DNA methyltransferase blocker RG108 protected the motor nerves nearly 100% after sustaining injury.

“It is an exciting first step,“ says Ratan. “I think this study has really implicated DNA methyltransferases in neurodegeneration in a way that no one has come close to."

Intriguingly, the Johns Hopkins team found that one of these DNA methyltransferases, Dnmt3A, was present in the right place – the nerve terminals – to potentially trigger ALS.  And, these enzymes could be found in these distal regions in mitochondria - potentially resulting in power outages, synaptic dieback and ultimately, the muscles to become unplugged.

“If we could actually show that ALS starts in the mitochondrial genomes at the nerve terminal,” says Martin, “that would be a new way of thinking.”

Now, Martin’s team is working hard to track down the genes in people with ALS that are silenced. Identifying these hypermethylated genes can give researchers a better idea of what signals could target the motor nerves for destruction, facilitating the design of better strategies to protect them in people with ALS.

But there may be no need to wait that long.  Several pre-cancer or cancer drugs that target methyltransferases are being developed or are already in the clinic. Emerging medicines that could be evaluated for the ability to slow or stop ALS.

The jury is still out whether such methylation-reducing medicines could be safe and selective enough to be useful to treat a neurodegenerative disease such as ALS.  But compared to potential medicines that target the histone deacetylases, there are considerable advantages to developing such a strategy for ALS according to Martin.

“If the changes occur at the nerve terminals at the neuromuscular junctions,” explains Martin, “we don’t have to worry about [the drug] crossing the blood brain barrier. That could not only be a new target but a new way of thinking about treating the disease.”

References

Chestnut, B.A., Chang, Q., Price, A., Lesuisse, C., Wong, M. and Martin, L.J.  (2011). Epigenetic regulation of motor neuron cell death through DNA methylation.  Journal of Neuroscience 31(46), 16619-16636. Abstract | Full Text  (Subscription Required)

Del Signore, S.J., Amante, D.J., Kim, J., Stack, E.C., Goodrich, S., Cormier, K., Smith, K., Cudkowicz, M.E. and Ferrante R.J. (2009) Combined riluzole and sodium phenylbutyrate therapy in transgenic amyotrophic lateral sclerosis mice.  Amyotrophic Lateral Sclerosis 10(2), 85-94. Abstract | Full Text  

Piepers, S.,et al.(2009) Randomized sequential trial of valproic acid in amyotrophic lateral sclerosis. Annals of Neurology 66(2), 227-234. Abstract | Full Text  (Subscription Required)

Rouaux, C., Panteleeva, I., René, F., Gonzalez de Aguilar, J.L., Echaniz-Laguna, A., Dupuis, L., Menger, Y., Boutillier, A.L. and Loeffler, J.P. (2007) Sodium valproate exerts neuroprotective effects in vivo through CREB-binding protein-dependent mechanisms but does not improve survival in an amyotrophic lateral sclerosis mouse model.  Journal of Neuroscience 27(21), 5535-5545. Abstract | Full Text

Ryu H  et al. (2003) Histone deacetylase inhibitors prevent oxidative neuronal death independent of expanded polyglutamine repeats via an Sp1-dependent pathway. Proceedings of the National Academy of Sciences 100(7), 4281–4286. Abstract | Full Text

Ryu, H et al. (2005) Sodium phenylbutyrate prolongs survival and regulates expression of anti-apoptotic genes in transgenic amyotrophic lateral sclerosis mice. Journal of Neurochemistry 93(5), 1087-1098. Abstract | Full Text  (Subscription Required)

Scott, S. et al. (2008) Design, power, and interpretation of studies in the standard murine model of ALS. Amyotrophic Lateral Sclerosis 9(1), 4-15.  Abstract | Full Text  (Subscription Required)

Steffan JS et al. (2001) Histone deacetylase inhibitors arrest polyglutamine-dependent neurodegeneration in Drosophila. Nature 413(6857), 739–743. Abstract | Full Text  (Subscription Required)

Sugai F, Yamamoto Y, Miyaguchi K, Zhou Z, Sumi H, Hamasaki T, Goto M, Sakoda S. (2004) Benefit of valproic acid in suppressing disease progression of ALS model mice. European Journal of Neuroscience 20(11), 3179-3183. Abstract | Full Text  (Subscription Required)

Van Lint, C., Emiliani, S. and Verdin E (1996). The expression of a small fraction of cellular genes is changed in response to histone hyperacetylation. Gene Expression 5(4–5), 245–253. Abstract | Full Text  (Not available online)

Wu X et al. (2008) Histone deacetylase inhibitors up-regulate astrocyte GDNF and BDNF gene transcription and protect dopaminergic neurons. International Journal of Neuropsychopharmacology 11(8):1123–1134. Abstract | Full Text

Yoo Y.E. and Ko C.P. (2011)  Treatment with trichostatin A initiated after disease onset delays disease progression and increases survival in a mouse model of amyotrophic lateral sclerosis. Experimental Neurology 231(1), 147-159.  Abstract | Full Text  (Subscription Required)

Further Reading 

Fischer, A., Sananbenesi, F., Mungenast, A. and Tsai, L.H.. (2010). Targeting the correct HDAC(s) to treat cognitive disorders.  Trends in Pharmacological Sciences 31(12), 605-617. Abstract | Full Text

Sleiman, S.F., Basso, M., Mahishi, L., Kozikowski, A.P., Donohoe, M.E., Langley, B. and Ratan, R.R. (2009). Putting the 'HAT' back on survival signalling: the promises and challenges of HDAC inhibition in the treatment of neurological conditions.  Expert Opinion Investigational Drugs 18(5), 573-584. Abstract | Full Text

Urdinguio, R.G., Sanchez-Mut, J.V. and Esteller, M (2009).  Epigenetic mechanisms in neurological diseases: genes, syndromes, and therapies. Lancet Neurology 8(11), 1056-1072. Abstract | Full Text  (Subscription Required)

Learn more about epigenetics and disease

Watch Epigenetics on PBS. See more from NOVA scienceNOW.

 

Going the distance. Motor neurons (green) innervate into the muscles of the growing mouse forelimb. Courtesy of Jeremy Dasen, PhD, New York University School of Medicine.

More than 70% of motor neurons in the spinal cord degenerate in people with ALS just one year after being diagnosed with the disease. To turn the tide of motor decline, researchers are developing stem cell-based strategies to regenerate the deteriorated motor nerves.

But some ALS experts worry that such strategies may simply take too much time to be effective.  Motor neurons could take by some estimates years to extend and plug back into muscles. The same time many people with ALS battle and ultimately die of the disease.

Researchers at Johns Hopkins School of Medicine introduced in 2008 another option. Healthy astrocytes are created in the cervical spinal cord to protect remaining motor neurons in the diaphragm from degeneration. The strategy, currently being developed in mice, is thought to work in part by reducing levels of glutamate in the central nervous system which at high levels can kill motor nerves.

Reporting in 2008, the team found that such a strategy reduced motor decline and increased survival of mice with an ALS-like disease 11%.

“We thought that it would be clinically more feasible to introduce astrocytes when compared to motor neurons and re-establishing all those neuronal connections,” explains Johns Hopkins School of Medicine neurologist Nicholas Maragakis MD. 

Now, the Johns Hopkins team reports that such a strategy using human astrocyte precursors appears to generate astrocytes in the cervical spinal cord but does not offer neuroprotection in ALS mice.

The study is published this month in PLoS One.

Scientists implanted healthy human astrocyte precursors into the cervical spinal cord of mice with an ALS-like disease.  The team found these cells developed into astrocytes.  But the scientists observed no difference in life expectancy.

 

Transplantation post-op. Researchers transplanted healthy human astrocyte precursors (green) which subsequently differentiated into astrocytes (red) in the spinal cord of mice with an ALS-like disease. Here, a section of the spinal cord is shown at end-stage. Adapted from Lepore, A.C. et al. (2011) PLoS One, 6(10): e25968.

The reason, suspects Maragakis, is that these transplanted cells simply did not have enough time to develop into astrocytes and protect the motor nerves from deteriorating.  The team reported as few as 50% of cells developed into astrocytes in mice at the end-stage of disease.

“Human cells mature much more slowly,” explains Maragakis.  “I think it just might take longer to get that [neuroprotective] effect.”

The team is now taking a look at mice with a more slowing progression form of an ALS-like disease to determine whether or not these strategies given enough time could protect the motor nerves from degeneration.

Looking ahead, the team hopes to develop transplantation strategies using astrocytes generated from the patients themselves to eliminate the need for immunosuppressive medications.  But the researchers caution that astrocytes created using existing reprogramming technologies may not be exactly the same as native astrocytes.  And even more importantly, re-introducing cells from people with ALS may instead contribute to the disease.

“It may be that a patient getting back his or her own cells may not be a good thing,” says Maragakis, “particularly for those with genetic forms of the disease.”

Meanwhile, researchers from BrainStorm Cell Therapeutics in Israel are developing a different strategy to prevent further deterioration of the motor nerves.  Astrocyte-like cells derived from patients’ bone marrow are genetically modified to produce substances including glial cell-derived neurotrophic factor (GDNF) which protect the motor nerves.  Called NurOwn, this strategy is currently being evaluated in Israel in an open label phase I/II trial.  First results are expected in early 2013.

References

Lepore, A.C., O'Donnell, J., Kim, A.S., Williams, T., Tuteja, A., Rao, M.S., Kelley, L.L., Campanelli, J.T., and Maragakis N.J. (2011) Human Glial-Restricted Progenitor Transplantation into Cervical Spinal Cord of the SOD1 Mouse Model of ALS. PLOS One, 6(10), e25968.  Abstract | Full Text

Lepore, A.C., Rauck, B., Dejea, C., Pardo, A.C., Rao, M.S., Rothstein, J.D., and Maragakis, N.J. (2008). Focal transplantation-based astrocyte replacement is neuroprotective in a model of motor neuron disease. Nature Neuroscience, 11(11), 1294-1301.  Abstract | Full Text

Further Reading

Maragakis N.J. (2010). Stem cells for the neurologist. Amyotrophic Lateral Sclerosis, 11(5), 417-423.  Abstract | Full Text (Subscription Required)