The 11th Annual ENCALS meeting highlights how TDP-43 spreads in MND

The European Network for a cure of ALS (ENCALS) held its 11^th Annual meeting in Sheffield from 31 May to the 2 June. The weekend was full of glorious British sunshine and more than 200 international scientists and clinicians were also able to enjoy a range of incredibly interesting talks about the latest developments in MND research.

A particular talk caught my attention on the first day by Dr Johannes Brettschneider from the University of Ulm in Germany. Dr Brettschneider explained how his research had shown the stages and spread of the protein TDP-43 in ALS (the commonest form of MND).

Dr Brian Dickie, Director of Research Development, said: “The key to defeating MND lies in fostering strong collaborations between neurologists, healthcare professionals, research scientists, early career investigators and students in the field of MND and the 11th Annual ENCALS meeting in Sheffield provided that opportunity. The MND Association was proud to support this event.”

‘Special’ staining

At the end of an afternoon of talks on the MND- causing genes C9orf72, FUS and SOD1, Dr Brettschneider engrossed over 200 delegates with his talk on the TDP-43 protein and how it spreads in ALS.

Although TDP-43 genetic mistakes are a rare cause of MND, scientists are especially interested in the TDP-43 protein because in the vast majority of cases of MND (irrespective of whether it was caused by an inherited genetic mistake), TDP-43 protein forms pathological clumps inside motor neurons.

The study (which is a collaboration between Dr. John Trojanowski and Dr. Virginia Lee from the Penn University Center of Neurodegenerative Disease Research in Philadelphia, America and the group of Dr. Heiko Braak in Ulm) used a technique known as ‘immunohistochemistry’.  This technique involves taking tissue samples of the brain and spinal cord from people who have died from ALS. The researchers would then make extremely thin slices of the tissue, which could then be stained using a ‘special stain’ and viewed under a microscope.

The stain used by Dr Brettschneider only ‘stained’ the TDP-43 protein in the samples, meaning that he could see the amount of TDP-43 in different areas of the brain and spinal cord.

Using the clinical information and TDP-43 staining this would allow Dr Brettschneider to stage the disease.

Image kindly provided by Dr Robin Highley, SITraN: (top left) a motor neurone with a skein-like neuronal cytoplasmic inclusion, next to a normal motor neurone (bottom left) on TDP-43 immunohistochemistry.

Axonal ‘telephone wires’ do more than just talking

Dr Brettschneider showed that TDP-43 increased in different areas of the brain and spinal cord during different stages of the disease. Amazingly, he also showed how ALS (characterized by clumps of TDP-43) spreads from one are of the body to another.

A motor neurone consists of three parts; the cell body, axon and nerve ending. The cell body contains the nucleus, or the control centre of the cell. When a message travels from the brain the cell body sends the message down the axon. Like telephone wires, the axon carries the message to the muscle, where the nerve endings cause the muscle to move.

However, in ALS it seems that these ‘telephone wires’ do more than just carry a message. The protein TDP-43 forms ‘clumps’ in the motor neurones and it seems that these clumps use the axon to travel from one motor neurone to the next (possibly explaining why someone get’s weakness in their arm and then their hand).

Another key finding was that TDP-43 clumps develop in the front part of the brain (prefrontal cortex), which is responsible for personality and may explain the development of cognitive symptoms.

Dr Brettschneider explained the importance of this research “While spreading of disease-related proteins has been described for other neurodegenerative diseases like Alzheimer’s disease or Parkinson’s disease, this had not been previously shown in ALS. Now, we can show evidence that supports a spreading of the major disease protein TDP-43 in ALS across specific regions of the brain and spinal cord with ongoing disease.

 If these findings can be confirmed (for example in cell culture or mouse model studies) then this could lead to the design of new treatments specifically aiming to impair the spread of TDP-43 protein clumps.

Dr Johannes Brettschneider from the University of Ulm in Germany at ENCALS

Furthermore, we believe that our findings offer a better understanding of disease progression in ALS.  Our data implies that TDP-43 spreads throughout the prefrontal cortex with ongoing disease, thereby lending support to the idea that all ALS patients could eventually develop “frontal type” cognitive deficits.”

The future

Dr Brettschneider commented why this research is important to people living with MND explaining that “If these stages can be reproduced in patients with ALS they could offer a new way to assess disease progression and response to new treatments. We hope that our study provides the essential groundwork for strategies designed to prevent pTDP-43 spread.”

This research is only the beginning and more work is needed, Dr Brettschneider also explained what he hoped to do next with these exciting results. “There were restrictions in time and availability of the tissue samples during this study, so we were unable to determine how and where exactly ALS begins in the very early stage of the disease. Therefore, an important next step in our work would be to analyze very early cases with ALS to look at TDP -43 spread as this offers the most promising window for therapeutic intervention.”

Reference

Brettschneider J, Del Tredici K, Toledo JB, Robinson JL, Irwin DJ, Grossman M, Suh E, Van Deerlin VM, Wood EM, Baek Y, Kwong L, Lee EB, Elman L, McCluskey L, Fang L, Feldengut S, Ludolph AC, Lee VM, Braak H, Trojanowski JQ. Stages of pTDP-43 pathology in amyotrophic lateral sclerosis. Ann Neurol. 2013 May 20. doi: 10.1002/ana.23937. [Epub ahead of print]

Anne Rowling Regenerative Neurology Symposium

The sun was (uncharacteristically!) shining on Edinburgh last week for a symposium to celebrate the launch of the new Anne Rowling Regenerative Neurology Clinic. The clinic, which opened to patients earlier this year, was founded following a donation by the author JK Rowling, in memory of her mother, who died from complications related to multiple sclerosis (MS).  Run by Professors Siddarthan Chandran and Charles ffench-Constant, the clinic aims to translate laboratory research into clinical trials for neurodegenerative diseases such as MS and MND.

The programme for the two-day meeting was packed with ‘big hitters’ from the world of neurology. In keeping with the regenerative neurology theme, the opening session was chaired by Sir John Gurdon, recent co-winner of the Nobel Prize for physiology and Medicine, whose pioneering work on cell cloning set the foundations for the more recent development of induced pluripotential stem cells, which are currently revolutionising medical research.

Different diseases, common challenges

The first day was given over to research areas such as multiple sclerosis, Parkinson’s disease and Alzheimer’s disease, as well as spinal injury and pain. What was also apparent is that different fields of neurology are wrestling with similar challenges: to diagnose disease earlier, ideally even before symptoms occur; to find biomarkers that tell us about the changes occurring in the Central Nervous System(CNS) at different stages of disease; to really understand the order in which these different aspects of pathology (the study and diagnosis of disease) occur and, given the theme of the conference, to sift the cellular changes caused by disease from the body’s attempts at cellular repair. All of these feed into the greatest challenge – how to take this accumulated knowledge from bench to bedside.

We can learn a lot from diseases that are further ahead in this process, such as the excellent overview by Prof Alastair Compston (Cambridge) on MS. It’s becoming clear that MS has distinct disease stages, starting off as an inflammatory disease, but progressing to a more ‘traditional’ neurodegenerative disease in more advanced stages. Whist there has been some considerable success in treating the former, the approaches to the latter have, as with MND, met with very limited success.

The use of imaging techniques to work out what is happening within the brain has been a vital factor in drug development for MS. As Prof David Miller (University College London) pointed out, magnetic resonance imaging (MRI) can pick up positive changes in small MS drug trials that are not large enough to show changes in disability. This sort of biomarker-based evidence gives drug companies the confidence to invest in the larger, much more expensive trials needed to show a clinical effect.

A presentation on the imaging of pain by Prof Irene Tracey (Oxford) provided a fascinating insight into the power of the placebo effect. She explained how neuroimaging has helped researchers to identify the brain regions associated with placebo effects and also gave examples of studies where the placebo effect has performed as well as (and even outperformed) commonly used painkillers! The power of placebo can be very strong indeed and it is important to always ensure that trials are rigorously performed to account for this.

Parkinson’s disease has always been viewed as a promising candidate for cell transplantation therapy, but clinical studies over the past 30 years have produced mixed results. Profs Roger Barker (Cambridge) and Anders Bjorklund (Lund University) discussed the various reasons for this ‘heterogeneity of response’ and how these are being addressed in the plans for a pan-European study.

In terms of cell transplantation, the approaches that will need to be taken for MND are very different from those for Parkinson’s disease. In Parkinson’s disease the strategy is to try and replace some of the key neurons that have died, but due to the immense length of human motor neurons, such a strategy of rewiring the nervous system is highly unlikely to work for MND. However, there are other approaches that can be taken, as Prof Clive Svendsen (Cedars-Sinai Medical Center) explained.

His approach involves a combination of gene therapy and stem cell therapy. By converting human stem cells into astrocytes, which are cells known to play an important role in keeping neurons healthy. By genetically modifying these cells to produce large quantities of a nerve protecting factor called glial-derived neurotrophic factor, and injecting them into the spinal cord of SOD1 rats, he has shown that the surviving motor neurons can be protected. He is in the process of gearing up for a phase 1 therapeutic trial in up to 18 carefully selected MND patients.

Disease in the dish

Prof Svendsen also briefly spoke about the promising research arising from the use of induced pluripotential stem cells (iPSCs) to study MND – a topic taken up in much more detail by Prof Jeff Rothstein (Johns Hopkins University) who highlighted recent advances in understanding the C9orf72 form of the disease.

It may be possible to create specially tailored gene therapy approaches for some forms of familial (inherited) MND, as is currently being attempted in SOD1 MND. Prof Rothstein’s initial work using iPSC-derived motor neurons suggests that this approach is also worth considering for the more common C9orf72 from as well.

Prof Steve Finkbeiner (University of California) who is collaborating in the Association-funded international stem cell initiative elaborated on the use of iPSCs as a tool for drug discovery, demonstrating how fully-automated robot-based systems can be used to follow the fate of thousands of individual human motor neurons in the dish over a prolonged time period. The great thing about robots is that they don’t need sleep, so can analyse the cells at all times of day and night. They do, however, have Twitter accounts, so they can report in to the centre staff when they have completed their experiments!

One of the exiting prospects of using these automated systems is the potential to screen thousands of compounds. If human motor neurons can be protected in the dish, there are no guarantees, but it at least shortens the odds that the human motor neurons can be protected in the human as well. There are still many improvements that can be made to the process, but screening work is underway, with a particular focus on drugs that stimulate cellular process called autophagy (a process in which a cell breaks down damaged components), which is believed to be protective across a number of neurodegenerative diseases.

There were many take home messages from this meeting, but what was abundantly clear from all the work presented was the enthusiasm of each speaker for their field of research and an optimism that we are on the cusp of major advances in understanding neurological conditions. Sharing of new knowledge across the various diseases and disciplines can only bring those advances closer.

Jolly Good Fellows!

There’s a scene in the 1969 film Battle of Britain where Laurence Olivier, who plays the Air Chief Marshal, is in a meeting with his two Vice Marshals. One of them complains that they don’t have enough planes; the other is more concerned with keeping the airfields working. Olivier silences them both by telling them that the fight will be won or lost on one key factor – the number of trained pilots.

It’s a rather cheesy film, but I used that story earlier this month to illustrate the importance of investing in bringing through the next generation of researchers in our battle to defeat MND.  We organised a ‘get together’ of our Lady Edith Wolfson Clinical Fellows at the Sheffield Institute for Translational Neuroscience (SITraN) to share their research findings with the donor who has so generously supported the scheme. The ‘get together’ also provided a wonderful opportunity for them to exchange information and expertise with each other, as well as all the staff of SITraN, who over the course of the day were frequently shuttling between the lecture room and their labs.

The Fellowships are aimed at attracting and training the brightest and the best Clinician-Scientists (or ‘Doctor-Doctors’ as I sometimes call them – with both a medical degree and a science PhD). Even so, I couldn’t resist using this cartoon in my introduction, although the reality is very different for our Fellows – the bar is set very high and even applicants for the Junior Fellowships need to have considerable research experience and be fully ‘lab tested’.

image courtesy of http://www.vadlo.com

Our host for the day was one of the world’s most respected MND ‘Doctor-Doctors’, SITraN Director Prof Pam Shaw, who welcomed everyone to the meeting and provided an overview of the multidisciplinary expertise and collaborative philosophy that underpins SITraN. Prof Shaw also has a great belief in the importance of nurturing the next generation of talent and it is no surprise that almost half of the Clinical Fellows in the programme are based at SITraN.

Are fit and active people more likely to develop MND?

Our first research presentation was from Dr Ceryl Harwood (Sheffield) who is carrying out research on the epidemiology of MND. Specifically, she is addressing the question of whether physical activity is a risk factor for MND. As she explained, this has been a long standing theory, showing us a quote from a medical journal written over 50 years ago which stated:

”Nothing has been said about the possible role in aetiology of a previous habit of athleticism. I have the uncomfortable feeling that a past history of unnecessary muscular movement carried out for no obvious reason may be followed in later life by the development of motor neurone disease in a statistically significant number of cases”

She outlined the plausibility that physical activity may contribute to a complex interplay between biological and genetic processes that may predispose an individual to develop the disease. Generating the evidence, however, is no easy matter, but she has developed and validated a novel questionnaire to measure physical history in adulthood, using data from a diabetes study in the 1990s where over 1,000 people had detailed measures taken of their actual energy expenditure.

A hundred of these participants have recently agreed to undergo rigorous face-to-face interviews and their responses were correlated with actual physical measures from over 15 years previously. In other words, she can now assess how accurately peoples’ recollection of their physical activity – both day to day work and vigorous exercise – links with their actual energy expenditure at the time. This questionnaire is now being used to compare the physical activity profiles in up to 350 people with MND and 700 control participants in Yorkshire and surrounding counties.

Should the results support the theory that physical activity is a predisposing factor in MND, she will be perfectly placed to delve into the genetic factors that underpin the selective vulnerability of motor neurons.

Repetition is bad….

Dr Pietro Fratta

Next up to the lectern was Dr Pietro Fratta, (University College London) who has been immersing himself in the mysteries of how the C9orf72 gene can cause neurodegeneration – especially MND and a related condition called Frontotemporal Dementia (FTD).

Like a needle on a vinyl record can sometimes stick and repeat the same fragment of music again and again, this gene sometimes carries a repeat in its genetic code – specifically with the letters GGGGCC occurring again and again.  Dr Fratta has examined many DNA samples from MND and FTD patients and finds that these ‘repeat expansions’ are very large indeed, occurring between 700 and 4000 times!

The process through which these repeat expansions cause nerves to die is still a mystery, but Dr Fratta showed results from his lab which suggests that rather than losing its normal function, the C9orf72 gene gains some additional activity, turning it into a ‘rogue’ gene. He and his colleagues have recently shown that the repeat expansions can lead to the formation of very stable chemical structures called G-quadruplexes that have been implicated in causing nerve damage in other disorders.

He is currently studying how these structures interact with other cellular components, interfering with normal neuronal function. He is also starting to look at possible therapeutic approaches in a collaboration with the UCL School of Pharmacy to develop compounds that will bind to and hopefully inactivate these structures.

Over lunch, we were given a guided tour of the superb SITraN labs by Prof Shaw. Although I strongly believe that research is only as good as the researchers doing the work, there’s no doubt that having a purpose built institute filled with state-of-the art technology certainly doesn’t do any harm!

Then it was back into the lecture room for our afternoon presenters.

A Sheffield double act

The post-lunch session was kicked off by Dr Robin Highley, a neuropathologist who has recently completed his Fellowship and now divides his time equally between pathology duties and MND research. Dr Highley’s area of expertise is in how neurons edit the genetic instructions into precise ‘blueprints’ to make proteins, the essential building blocks of every cell in our body.

He used an entertaining analogy of making dresses form a pattern to describe the process of how DNA is made into RNA copies which can be ‘tailored’ into slightly different protein designs (to find out more about how DNA makes RNA and subsequently proteins see our earlier blog post).

Dr Johnathan Cooper-Knock, MRC/MND Association Lady Edith Wolfson Clinical Research Fellow

Using a variety of approaches he has looked at gene expression (which genes are being switched on and off) and gene splicing (how the RNA copies are edited) patterns in both inherited and non-inherited MND, as well as in non-MND states. He finds changes occurring in thousands of genes, but by performing searches on databases of the ‘function’ of each gene he can then sort them into different groups (which are then involved in key cellular processes). This provides important clues as to which cellular pathways are altered in MND, which will help researchers around the world to focus their attention on the most common changes and hopefully start addressing the question of how these may be slowed or stopped.

Dr Highley focused his talk mainly on the TDP-43 and SOD1 forms of inherited MND, with his colleague and fellow ‘Fellow’(!) Dr Johnathan Cooper-Knock, concentrating on the C9orf72 form (the most common cause of inherited MND). Through the MND Association’s DNA Bank  he has been able to obtain a large number of cell lines from patients with C9orf72 MND, along with detailed clinical information, which will allow him to compare patterns between those with fast progressing and those with more slowly progressing disease.

Although at a much earlier stage in his research, having started only 6 months ago, Dr Cooper-Knock has already identified some specific gene expression effects that may be distinct to the C9orf72 form of the disease. For more details about Dr Cooper-Knock’s work see our earlier blog post about his fellowship.

BioMOx and beyond

It was fitting that Dr Martin Turner (Oxford) gave the closing presentation. Not only was Dr Turner the first recipient of a Lady Edith Wolfson Fellowship, but he has recently been awarded a new five-year Senior Clinical Fellowship through the programme – these are highly prestigious awards, with only one in seven applicants successful.

Dr Martin Turner, MRC/MND Association Lady Edith Wolfson Clinical Research Fellow

Titling his talk ‘BioMOx and beyond’ Dr Turner outlined the challenge of identifying a specific signature of MND. He showed that whilst there is unlikely to be a single test for MND, a combination of tests (involving brain scanning and eye tracking techniques together with chemical analysis of blood, urine or cerebrospinal fluid) are showing some promise in aiding and speeding up the diagnosis, as well as predicting how the disease is likely to progress within an individual.

He highlighted the importance of international collaboration, such as the new formal link with Dr Mike Benatar in Miami, who for several years has been studying people at risk of developing inherited MND. Indeed, Dr Turner apologised for missing the morning speakers at Sheffield as he had been busy with one of Dr Benatar’s study participants in his MRI scanner at Oxford!

On the subject of international collaboration, our most recent Clinical Fellow, Dr Jemeen Sreedharan, was unfortunately unable to attend as the first two years of his Fellowship is based at the University of Massachusetts, returning to the University of Cambridge to complete his research. We look forward to having him at the next Fellows get-together!

Major new research finding raises some old questions

Hot on the heels of ‘Brain Awareness Week’, comes ‘National Science Week’, with the University of Sheffield enthusiastically organising a numerous activities in their week-long Festival of Science and Engineering, including today’s Open Day at the Sheffield Institute of Translational Neuroscience (SITraN). This event was to include talks by Dr Chris McDermott and Prof Pam Shaw on MND and the role that SITraN plays in the search for effective treatments for neurodegenerative disease.

Unfortunately…… last night’s snow has put the kibosh on that, so instead of heading up the M1 to Sheffield, I decided to use some of the saved time to catch up on some reading – in particular a recent paper that came out in the journal Nature, from an international consortium, led by the scientists from the Austrian Academy of Sciences in Vienna.

The best seven pages in ten years

A senior MND researcher emailed me to say it’s one of the best papers he’s read in the past 10 years and I can understand where he’s coming from. Not only does the research identify a previously unknown cellular process that causes selective motor neuron degeneration, but it also appears to tie together several of the pieces of the pathological jigsaw: disruption of RNA metabolism, oxidative stress and programmed cell death pathways.

As impressive is the sheer amount of work that has gone into this seven-page paper. OK, there are also several extra pages of online supplementary material (one of the great benefits of online publication) but I reckon there is the equivalent of at least three PhD theses and several years of work in there!  

In a nutshell, the researchers created a mouse that has a defect in an enzyme called CLP1 and these mice develop progressive motor neuron degeneration. I’m not going to go into the detail, but rather focus on one interesting item that was buried in the text.

Genetic environment matters for CLP1

When researchers initially tried to create the mice they found that the mice all died well before birth. So they tried using a different strain of mice, but got the same result.

A third strain produced live mice, with normal numbers of motor neurons at birth. However from about the age of four months, these mice then developed a progressive muscle weakness and loss of motor neurons over the course of several months.

The paper focuses in on what’s going on in these mice, but it also raised additional questions for me, such as:

“Why did these mice survive into adulthood, when two other mouse strains didn’t – and is there something different in the genetic make-up of these mice that has basically protected them into adulthood rather than killing them as embryos?”

MND Association funded research on genetic environment

Other groups have noticed that when SOD1 mice are bred on different background strains, it can have a profound effect on disease progression and survival. This brings us nicely back to SITraN, as Prof Shaw and her colleagues are looking at precisely this issue, in an MND Association-funded collaboration with Prof Caterina Bendotti in Milan.

They are looking at the gene expression profiles (basically which genes are switched on and off) in the motor neurons of two strains of SOD1 mice, one of which develops the disease and later age and also lives much longer.  By working out patterns that are linked to specific biological processes, they are starting to pinpoint pathways which are driving the disease and also which ones might be slowing the disease. Some of their findings were presented at the most recent International Symposium (Abstract C61).

If there are protective genes at work in the mice, might the same be happening in humans?

The search for ‘good’ genes hots up

I’m often asked about Steven Hawking – how come he’s lived so long?  For years, one of my pet theories has been that there is something in his genetic make-up that didn’t stop the disease from occurring, but is ‘pushing back’. That’s becoming an increasingly popular and productive area of investigation – as genetic researchers extend their focus from finding ‘bad’ genes that cause or predispose people to develop MND, to potentially ‘good’ genes that might slow down the disease. A couple of candidates have been identified, most notably the EphA4 gene.

The search for these disease-modifying genes needs joined up collaboration between researchers around to world and it’s heartening to see how everyone in the field is starting to get together to pool their samples and data, which will allow the genetic profiles of those with exceptionally slowly progressing MND to be analysed in much larger numbers than ever before. If good genes can be identified and their roles understood, it will open up exciting new treatment opportunities.

Vive la difference!

It could be easy to assume that one motor neurone is pretty much like another, but a series of presentations on Thursday clearly showed that we need to be a little more sophisticated in our thinking.

Dr Hynek Wichterle (Columbia University) discussed how in the developing embryo, there are distinct regional subtypes of motor neurones in the spinal cord. For example, the motor neurones at the bottom of the spine can be easily distinguished from those at the top of the spine by the pattern of genes that are switched on and off – in a nutshell, different motor neurones have their own regional ‘postcode’.

Dr Wichterle went on to show that you can generate motor neurones with specific postcodes  in the lab, from embryonic stem cells . Having motor neurones  that  hopefully reflect different aspects of MND will allow researchers to better understand the subtleties of motor neurone function and develop more relevant treatment approaches.

It’s well known that some motor neurones are particularly resistant to the disease, such as the so called ‘oculomotor neurones’ that control eye movements. By looking at  the pattern of genes  being switched on and off in early mouse embryonic development, Dr Wichterle was able to induce the same pattern in mouse stem cells to create what look very much like oculomotor neurones in the dish. There is some further work to be done to check that they are indeed what he thinks they are, but they could be a very important tool in helping to understand why some motor neurones are tougher than others.

Continuing the theme, Dr Georg Haase  (Marseille) introduced an elegant way of separating different populations of motor neurones from mouse spinal cord. He focused on two subtypes of motor neurone – those that connect to the limb muscles  and those that connect to the main trunk of the body. After they are separated out, they can be grown in culture (in a dish in the lab) . He then showed that these two different subtypes of motor neurone acted very differently in their response to chemicals known as neurotrophic factors . The word neurotrophic can be literally translated as ‘nerve nourishing’ and these chemical compounds have attracted a lot of attention in the past as possible therapies for MND, with a couple being tested in large clinical trials . Unfortunately these trials have not shown any effect, but this could be because  – as Dr Haase showed in his lab studies – each neurotrophic factor only acts on a proportion of motor neurones. He suggested that the different ‘survival profiles’ he sees in his cellular studies  provide a rationale for selecting combinations of different neurotrophic factors for further testing in mice – and hopefully in man.

Prof Pam Shaw (University of Sheffield) also asked the question of what makes  oculomotor neurones  different, but her presentation took us from mouse to man. Using a technique called laser capture microdissection, which allows individual motor neurones to be removed from post mortem spinal cord tissue form MND patients, she and her colleagues are able to examine which genes  were switched on and off in the cells. Research like this requires very high quality tissue, removed shortly after death, but Sheffield happens to host one of the best MND tissue banks in the country.

She compared these ‘gene expression patterns’ in the disease-resistant oculomotor neurones with the more vulnerable motor neurones  from the lower spinal cord. There were  considerable differences in the patterns, with a much larger number of genes being switched on in the oculomotor neruones. Many of these genes could be dividied into functional families, which act on a similar biological pathway. Key protective processes activated included neurotransmission, mitochondrial function and proteasomal function – all of which are strongly implicated in MND. If human oculomotor neurones  survive by gearing up these  protective pathways, it provides possibilities that drugs can be found which act in the same way.

Our International Symposium website news stories:

International Symposium closes in Chicago

International Symposium focuses on clinical trials

International Symposium focuses on carer and family support

International Symposium begins in Chicago

Researchers unite at our International Symposium on MND

After you’ve finished reading the symposium articles that interest you, we’d be grateful if you could spare a few minutes to fill in our short online survey on our symposium reporting. Your comments really are useful and allow us to continually improve our symposium reporting. surveymonkey.com/s/alssymp 

Mastering Pac-Man

Growing up in a seaside town in the 1980s led to me spending a lot of my “formative years” in the local amusement arcades, playing iconic video games like Space Invaders (at which I was average) and Asteroids  (I was the kid to beat!). One game I never got the hang of was Pac Man, where you had to guide a munching yellow ball around a maze eating up lots of dots. I even bought a book ‘Mastering Pac Man’ which didn’t help much – I was just plain rubbish!

Dr Brian Dickie and Pac Man

Talking about rubbish, fast forward 30 years and Prof Anne Simondsen from The University of Oslo is on the platform in Chicago describing the ways in which neurones deal with their cellular rubbish, such as poorly manufactured or damaged proteins.

Much as the dots are gobbled up by Pac Man, so cells employ a couple of basic ways to dispose of and recycle damaged proteins. One is a process called proteasomal degradation, which has been discussed on this website before by one of our guest bloggers, Prof John Mayer. However, some proteins, especially protein aggregates, are a bit too large and tough to be broken down by the proteasome, but they can be degraded by another process called autophagy – which can literally be translated as ‘self-eating’.

Prof Simondsen explained how aggregated proteins and even larger cellular structures can be packaged up for destruction by being encapsulated within membranes, forming structures called autophagosomes. These in turn seek out a structure called the lysosome, which contains digestive enzymes which break down the protein rubbish and keep the cell ‘spick and span’.

Using studies involving fruit fly models (a favourite model for cell biologists) she showed that the autophagy process declines with age, which is unfortunate because protein damage tends to increase with age. It was therefore no surprise that the level of autophagy in the brain was directly associated with the accumulation of protein ‘inclusions’, the health of neurones and the life expectancy of the flies.

Much of the evidence supporting a role for autophagy in neurodegeneration comes from the field of Huntington’s disease. Prof Simondsen explained how a particular mutated protein – called huntingdin – accumulates in the dying neurones and is a classic pathological hallmark of the disease. She demonstrated that autophagy normally plays a central role in the disposal of damaged huntingdin, but the system simply cannot cope with the amount of damaged protein, which starts to accumulate and literally ‘gunge up’ the cell. However, by stimulating the manufacture of additional autophagy machinery, the amount of aggregated protein can be reduced, helping to protect the neurones. This has so far only been shown in simple lab models of Huntington’s disease, but the encouraging results mean that further development to try and develop a treatment is underway.

So, could a similar approach be useful in other neurodegenerative diseases such as MND? Certainly, for some types of MND, such as those linked to a rare gene mutation called CHMP2B, it appears that a component of the autophagic machinery is impaired, which leads to protein build-up in cell models.

Dr Eiichi Tokada (Umea University, Sweden) provided further evidence that autophagy plays a role in disease progression in the SOD1 form of MND. By switching off a crucial component of the autophagy machinery, the disease progression in SOD1 mice was accelerated and their lifespan shortened. In addition, when he examined the spinal cords of the mice, he saw an increased presence of the characteristic SOD1 protein aggregates – a pathological hallmark of the disease.

Dr Faisal Fecto (Northwestern University, USA) provided evidence from a third genetic cause of MND. He showed how mutations in the Ubiquilin 2 gene disrupt the autophagy pathway by stopping the autophagosomes from linking up with the lysosomes.

So, it certainly seems that autophagy has a role to play in some of the rarer familial froms of MND. It remains to be seen to what extent it is involved in MND more generally, but it may mean that the potential treatment strategies being developed for Huntington’s disease may offer future opportunities for MND as well.

Our International Symposium website news stories:

International Symposium closes in Chicago

International Symposium focuses on clinical trials

International Symposium focuses on carer and family support

International Symposium begins in Chicago

Researchers unite at our International Symposium on MND

After you’ve finished reading the symposium articles that interest you, we’d be grateful if you could spare a few minutes to fill in our short online survey on our symposium reporting. Your comments really are useful and allow us to continually improve our symposium reporting. surveymonkey.com/s/alssymp

University of Bath research shows how angiogenin affects motor neurone survival

Published in the prestigious journal Nature Communications and funded by the Wellcome Trust, University of Bath researchers looked closely at the structure of 11 mutated angiogenin proteins, and how changes in their structure influenced activity, function and survival in motor neurones.

Dr Brian Dickie, Director of Research at the Association, said “The researchers at the University of Bath have skilfully combined aspects of biology, chemistry and physics to answer some fundamental questions on how angiogenin can damage motor neurones. It not only advances our understanding of the disease, but may also give rise to new ideas on treatment development.”

Mutated forms of the gene editing protein, angiogenin, are known to cause MND in some families affected by the inherited form of the disease. This is due to a mutation in the normal angiogenin gene (a mistake in the instructions for making the protein) that leads to the production of a faulty/mutated angiogenin protein.

This new research expands on our knowledge of angiogenin, as back in June this year angiogenin, was identified by Irish researchers as playing a vital role in motor neurones with the effects tested in a mouse model of MND.

Creating images with x-rays

University of Bath researchers used a technique known as x-ray crystallography (a method that uses x-ray light to produce highly detailed images of the structure of a protein). Using this technique they were able to view the structure of angiogenin, and 11 angiogenin MND mutants, in exquisite detail. Specifically the researchers looked at the active site (where all the action happens) for changes that may affect the function and activity of the mutated proteins.

These x-ray crystallography images show stunning structural differences in the angiogenin MND mutant proteins, particularly around the active site, and the researchers found that these structural changes caused the mutated angiogenin proteins to have varying levels of activity.

Dr Vasanta Subramanian, who was involved in the research, said: “We hope that the scientific community can use this knowledge to help design more drugs that will bind selectively to the defective protein to protect the body from its damaging effects.”

An MND Association funded study has also used this technique of x-ray crystallography to look at the structure of mutant SOD1 proteins, which are another cause of inherited MND. Under the leadership of Prof Samar Hasnain, Dr Neil Kershaw is also one step ahead and is examining potential drugs to ‘mend’ these abnormal SOD1 proteins. This shows how this type of research, investigating structure and activity of the mutated proteins, can influence the development of new drugs.

What happens to angiogenin in the cell?

As well as changes in the activity of the protein the structural changes cause the angiogenin MND mutants to lose their function, leaving a protein sitting around in the cell, unable to do anything! The researchers found that it is essential that angiogenin is transported from the cytoplasm to the nucleus of the cell in order for the protein to function properly. This is an important step during the process in which proteins are made, if this does not occur then the protein will not function properly. In the angiogenin MND mutant proteins this does not occur correctly with either none, or very little, angiogenin being transported to the cell nucleus. This means that the angiogenin MND mutant proteins do not function properly compared to the normal angiogenin protein.

Stress Granules

The University of Bath researchers also found that the angiogenin MND mutant proteins prevented the motor neurones from producing ‘stress granules’. These are little packages that appear when a neurone is under stress, such as when low oxygen levels occur. When this stress factor occurs they release their contents to protect the motor neurone from damage.

These stress granules are the motor neurones natural defence mechanism. Without these granules the cell’s natural defences are lost and this influences the survival of the cell.

In Summary

This research gives us an important insight into how MND mutations affect the normal structure and function of angiogenin in motor neurones in some inherited forms of the disease. This not only advances our understanding of the disease, but may give rise to new ideas on treatment development in the future.

University of Bath press release

Journal article

Research we fund

New fellowship explores how C9ORF72 causes MND

Dr Johnathan Cooper-Knock, MRC/MND Association Lady Edith Wolfson Clinical Research Fellow

Dr Johnathan Cooper-Knock from the Sheffield Institute for Translational Neuroscience (SITraN) has been awarded with the fifth Medical Research Council (MRC)/MND Association Lady Edith Wolfson Clinical Research Fellowship.

Through his three-year fellowship, Dr Cooper-Knock will use the MND Association’s DNA bank to study how recently discovered mistakes (known as mutations) in a gene called C9ORF72 can cause the disease.

Dr Johnathan Cooper Knock explains, “I believe that the genetics of MND are a key to understanding both the cause of the disease and how to treat it. The discovery of mutations in C9ORF72 are a great opportunity to get a hold on mechanisms of disease which has so far been elusive. I am excited by the opportunity my fellowship will give me to pursue this important discovery.

“By the end of my fellowship I aim to have contributed significantly to the understanding of disease mechanisms related to C9ORF72 dysfunction in MND. As a result I hope to have identified a number of therapeutic targets for development into new treatments by myself and others.”

C9ORF72: the facts so far

We know that a repeated six-letter code within a gene called C9ORF72 can cause MND and a related condition called fronto-temporal dementia (FTD) for approximately 40% of cases with a positive family history of MND and/or FTD.

Most genetic mistakes found in MND to date have been swaps of genetic letters, which can change the meaning of that part of the gene. The C9ORF72 genetic mistake on the other hand, is a repeat expansion. This means that six letters within the genetic code (CCCCGG) are repeated hundreds of times for people with C9ORF72 MND. In healthy individuals, this repeat is found about 30 times. We already know that the exact size of the repeat varies substantially between people with this genetic mistake. How this repeat causes MND and how the size of the repeat may affect disease progression is currently unknown but this is something that Dr Cooper-Knock wants to find out.

We also don’t know what role C9ORF72 normally has in the body. Even its name, which stands for ‘chromosome 9 open reading frame 72’ refers to where it is in the genetic code and not what it does. This isn’t unusual as it’s currently estimated that we have over 20,000 genes, and understandably, researchers haven’t yet found out what every one of these does – including C9ORF72.

So far, 96 journal articles have been published about C9ORF72 (by searching on Pubmed for C9ORF72). The oldest of these was published in 2011, and describes the original MND/FTD C9ORF72 finding. All subsequent articles on C9ORF72 have been of a direct consequence from this pivotal genetic discovery in the past year.

These 96 studies were focused on finding out how many people have the C9ORF72 genetic repeat and finding out what this mistake ‘looks like’ both clinically in terms of progression rates, age of onset and symptoms; and in terms of post-mortem findings to compare with other forms of MND. Coincidently, the most recent post-mortem and clinical C9ORF72 finding was authored by Dr Cooper-Knock (when searching for C9ORF72 and post mortem on PubMed).

It’s reassuring to know that researchers aren’t resting on their laurels with this genetic finding. There’s a huge international research effort in place to push forward our understanding of C9ORF72, with a number of our own newly funded projects, starting later this year, dedicated to creating new laboratory models of this genetic mistake to better understand how it can cause the disease.

How do we currently think C9ORF72 causes MND?

Due to the sheer size of the repeat expansion in C9ORF72, it’s thought that it causes MND by disruption of the editing process of genetic information.

I’ll explain: In real life terms, our DNA can be thought of as being held within a library, which is the control centre of the cell (the nucleus). Each book (gene) is stored on a particular shelf (chromosome). Gene ‘books’ aren’t allowed to be taken out of the nucleus, but they can be photocopied. These copies (RNA) are edited and transported out of the nucleus to be used as instructions to create proteins that perform specific roles in and sometimes out of the cell.

Unlike real life books, genes are fraught with errors, variations and nonsense from one person to the next. It looks messy, but it’s normal. Genetic editors are needed to edit and chop the RNA into a readable format so that it can be understood by the parts of the cell that use RNA as instructions.

As normal, healthy copies of C9ORF72 hold approximately 30 repeats to be chopped out as RNA, the effect of having much larger repeats may be having a knock-on effect on the efficiency of the editing process. This could then lead to a much higher risk of developing MND.

Finding out exactly how C9ORF72 can cause MND, and whether this theory is right, will provide us with a deeper insight into MND and potentially provide therapeutic targets that can be further investigated.

Dr Cooper-Knock’s fellowship

Dr Cooper-Knock will be using a cutting-edge genetic technique called ‘gene expression profiling’ to study the various levels of RNA in samples provided by people with the C9ORF72 genetic mistake. From this, he’ll find out which genes are switched on and off because of the C9ORF72 repeat expansion.

He will also study whether the size of the C9ORF72 repeat expansion has an effect on symptoms or progression rates to identify factors that may modify disease progression and may therefore be targets for future therapies.

Technology specialised for identifying misassembled RNA will also be applied to skin cells donated by people with the C9ORF72 repeat expansion who have MND/FTD and healthy controls. This will help to elucidate what the C9ORF72 protein does.

As well as skin cells from people with MND/FTD, this study will use post-mortem brain and spinal cord tissue from people with the C9ORF72 repeat expansion and healthy controls within the Sheffield tissue bank; as well as cells from the blood of C9ORF72 patients and healthy controls obtained from the MND Association’s DNA Bank.

Talking about the importance of people with MND having provided these numerous samples Dr Cooper-Knock said “Without the participation of patients and their families MND research will get nowhere; and equally with their participation, doors are opened towards new and exciting treatments. At this time, with discoveries like the mutations in C9ORF72 to build from, we can do even more with the participation of those who have been affected by this disease, who like us are passionate to see it cured.”

More information:
Read our official news release on our MND Association website.
Find out more about our DNA bank.

Discussing MND in Dublin

Delegates to last weekend’s ENCALS (European Network for the Cure of ALS) meeting in Dublin were met with uncharacteristic hot and sunny weather – enjoyed by the numerous Stag and Hen parties wandering the city centre, but not by the 200 people ensconced in the impressive, new Biomedical Sciences Institute at Trinity College, from 8am to 7pm, for a packed programme of presentations and debate.

ENCALS was established to help develop the standards of clinical and biomedical MND research across Europe and create a more collaborative environment for researchers, industry, funding agencies and Patient Associations. However, the meeting had a very transatlantic flavour, thanks to the participation of several of the leading researchers from North America.

With around 40 speakers, as well as numerous poster presentations, there is too much to cover in a few hundred words, so I’ll focus on just a few of the key themes that were covered. I also apologise for the quite technical language, which may make for hard reading, but is a positive in that it reflects the increasing complexity and sophistication of MND research.

Can we block the ‘molecular funnel’?

The opening speaker, Prof Teepu Siddique, from Northwestern University in Chicago, spoke on The molecular funnel of neurodegeneration. His view of MND is that it may have a large number of different causes, but the way a motor neurone dies will probably be similar, no matter what the original cause. We’re currently finding lots of new genetic factors involved in the disease, but we don’t understand how many of these genes work in health, much less how they malfunction in disease. So, the mouth of our funnel is getting wider.

Prof Siddique’s view is that by focusing on the cellular changes that are common to all forms of the disease, it gives us possible therapeutic targets that could be relevant to all forms of MND. It’s easier to block the funnel at its narrowest point.

He discussed how the degradation of incorrectly formed or damaged proteins is a classic hallmark of all forms of MND. While the way in which the proteins are damaged may differ from one form of MND to the next, it’s the cell’s inability to correctly deal with these proteins that may be a good target. If we can normalise or improve this process, it may keep the motor neurones functioning for longer.

Prof Orla Hardiman, the meeting organiser from Dublin, discussed the need for much larger and more detailed study of large numbers of patients, to attempt to unpick the environmental influences that undoubtedly exist.

A question that many people often ask is whether MND is occuring more often in younger people that in the past. Intriguingly, Prof Hardiman’s ‘population-based’ research using the Irish MND Register suggests the opposite – the average age of symptom onset is getting older. She suggests that continued improvement in medicine and diet means that the population in general is healthier, so our ‘biological age’ is slowing. If age-related diseases such as MND are linked to ‘biological age’ rather than ‘actual age’, it would explain this surprising trend.

Good Genes/Bad Genes

While factors that cause or predispose towards MND are clearly the subject of intensive research, there is of course also interest in factors that might prevent or slow the disease. Some of these potentially ‘good’ genetic variants are being explored:

• Prof Wim Robberecht’s group (University of Leuven) is examining the function of a gene called ephA4, which appears to correlate with longer survival in humans. This work is supported by studies in zebrafish and mouse models of MND.
• Prof Kevin Talbot (University of Oxford) showed data that suggests that by increasing activity of a gene called smn1 might be beneficial to motor neurones. This is a strategy that is being followed for a predominately childhood motor neurone disease called Spinal Muscular Atrophy, so if these approaches work in this particular condition, they might be of benefit in other, adult onset motor neurone diseases.
• Prof Robert Brown (University of Massachusetts) presented early data from a study of a variant in a gene called sarn1, which appears to protect motor neurones from damage….at least in fruit flies and mice. Work is ongoing to see whether it also has relevance in humans.

In contrast, Dr Andrea Calvo (University of Torino) provided information from Italian patients confirming studies in other populations that a variation in the unc13A gene can speed up disease progression.  However, the important issue about these disease-modifying genes – and it doesn’t matter whether they speed up or slow down MND – is that they all represent potential therapeutic targets.

Not just about the motor neurones!

We know that motor neurones do not die alone. Other parts of the brain and spine can be affected, but it’s the motor neurones that ‘bear the brunt’. 

Dr Sharon Abrahams (University of Edinburgh) provided an excellent overview of the range of cognitive and behavioural changes that can occur in the disease, indicating damage to other part of the brain, in particular the frontal lobe. Thankfully, the ‘real world’ effects of frontal lobe changes are usually subtle, but the fact that they can be picked up by psychological tests and MRI scans will help in defining specific ‘subtypes’ of MND which may require additional approaches to managing the disease.

Dr Martin Turner (University of Oxford) outlined evidence from a number of clinical research studies, including his own that nerve cells, called interneurones, might be involved early in the disease. These particular neurones usually play a role in calming down motor neurones, so if they are damaged or lost, the motor neurones themselves become over-excited and stressed, which leads ultimately to their degeneration.

Dr Turner’s evidence comes mainly from clinical imaging and electrophysiology studies in MND patients, but his theory was supported by a presentation from Dr Tennore Ramesh (University of Sheffield) who works with zebrafish models of MND. He showed results using zebrafish that carry a human SOD1 gene known to cause MND. The fish develop a form of MND in adulthood, but the very earliest signs of nerve damage actually occurs in specific types of interneurones that connect with the motor neurones, with the motor neurone damage occurring much later, closer to the onset of symptoms.

Presentations also covered the role of non-neuronal support cells, such as microglia and astrocytes, both of which have been the subject of extensive research in recent years, as they appear to play a role in the speed of progression of the disease. Prof Jeff Rothstein (Johns Hopkins University) introduced a new cellular player to the MND field, called the oligodendrocyte. These specialised cells have been known for many years to play a role in helping neurones to carry electrical signals, as well as helping them to maintain energy levels. Although they are known to be involved in multiple sclerosis, they hadn’t attracted much attention in MND.

Prof Rothstein showed that in human post mortem MND brain tissue, there is evidence that the brain has been making oligodenrocytes. This is certainly very clear in SOD1 mice, where  a massive production of new oligodendrocytes occurs. However the total number of these cells was not increased in the mice, suggesting that older oligodendrocytes were being killed and getting replaced.

He suggested that the new ‘immature’ oligodendrocytes are not nearly as efficient in their supporting role, especially when it comes to supporting motor neurones in maintaining their energy balance. This provides two possible treatment approaches – either try to keep the existing oligodendrocytes healthier or find a way of making sure that their replacements reach their full functional maturity.

I’ve no doubt we’ll be hearing a lot more about these cells in the future.

‘Hothouse’ meeting on drug discovery

Sadie’s recent post on the emerging partnership between Peakdale Molecular and the Sheffield Institute for Translational Neuroscience prompted me to say a little about last month’s Drug Discovery Workshop, held in Washington DC and organised our friends at the ALS Association.

I was fortunate to be invited along to this workshop, which brought together over 100 representatives from industry, academia, drug regulators and government and charitable funding agencies, to share their findings and discuss the future directions and opportunities for MND drug discovery. ‘Hothouse’ meetings like these are vital in giving those working in academic labs an important insight into the complexities of turning new knowledge of disease processes into ‘druggable’ compounds.

Those from industry get to see the new theories that are coming out of the academic labs, while the funders can start to identify where targeted early-stage support may help to encourage industry to follow up with the larger-scale investment needed to take ideas from bench to bedside At a time when some of the biggest drug companies are pulling back from working in neurodegeneration, the mood at the meeting might have been muted, but delegates were positively upbeat. One cause for optimism is that some companies, such as Biogen Idec, have seized the opportunity to fill the gap and increase their investment in this area.

Moreover, universities around the world have benefitted from an influx of new staff with extensive expertise in drug discovery, strengthening one of their historical areas of weaknesses. Universities are generally very good at unpicking the complex biological processes that occur in health and disease, but very poor at turning this knowledge into treatments. Another reason for the optimistic mood at the meeting was the clutch of new gene discoveries that occurred last year, in particular the identification of the chromosome 9 form of MND  which promises to open up many of new secrets of the disease. Researchers have collectively now found about two-thirds of all the causes of familial MND. As we identify more causes, generate better models and home in on the common cellular changes that drive the disease, the opportunities for drug development are going to increase.

Meetings such as this help focus attention on the major challenges – but also the exciting opportunities - that lie ahead.