Wednesday, 29 July 2015
Dr Richard Mead Kenneth Snowman and MND Association Lecturer in Translational Neuroscience
Research interests: Drug discovery in motor neurone disease (MND) from target identification, through in vitro screening to in vivo disease model testing.
Why do we still use animals in scientific research? Can we not
replicate in vivo (in whole body) work without animals?
I think the main justification for that is that we can’t replicate what happens in a whole animal in a tissue culture dish, or in a computer model. If we take, for example, the motor system, there’s a very complex interplay between, upper motor neurons and lower motor neurons and then the muscles that they connect to. There’s also astrocytes, glial cells and microglial cells which all interact within the central nervous system. People are working towards 3D co-cultures but these are composed of two or three components of something that’s way more complicated than that.
The other thing to consider, and this is true of any model of disease, is that you still have to confirm any discoveries in whatever you are modelling. So if you were modelling what happens in an animal using a tissue culture dish or a computer, you would still have to go back to the original (animal) model to confirm the findings.
Hopefully one day, we will be able to replace all animal use with other models, but we’re not quite there yet.
If a drug is found to work in an animal model of disease, why does it not always work in humans?
People tend to blame the animal model, suggesting that the model isn’t representative of the human disease and at a fundamental level this is true. We have to consider that all models have their limitations.
A British mathematician George E. P. Box said, ‘All models are wrong, but some models are useful’, and this applies to the models used in science. So the key is to work out whether your model is useful and how. One goal of SITraN is to be experts in the models so we understand their limitations. We also develop new models that can complement the ones we already have. This allows us to look for positive results in different types of model, increasing our confidence in them.
Personally I think the bigger issue is ‘translating’ mouse results to human studies and again we try and do this in a very rigorous way in SITraN. There are many steps to progress from a positive result in a mouse study, to a positive result in a clinical trial. If a drug is found to work in a mouse, it is very hard to work out an equivalent dose in humans, this can be estimated by weight, or (more accurately) body surface area. This is based on the fact that the smaller an animal, the faster their metabolism, and therefore, the faster they get rid of a drug from their body. We can also measure the levels of the drug in our mouse models and in humans and try and match them. An even better method is to use a ‘biomarker’ (a biological indicator e.g. expression levels of a gene) to show us how much of a drug has reached the relevant tissue in the mice, then measure the same thing in patients. We are working on measures that can be used in both mouse models and in humans to measure drug effects or the disease process.
Finally, it is important to make sure clinical trials have enough participants to detect a similar effect as was seen in the mouse model, and this isn’t always the case. So all in all, the process of translating treatments through from mouse to human is very problematic.
Can you give some examples of when animal work has helped to develop effective treatments in human disease?
Almost every disease that you can think of (which has a treatment). For example, diabetes used to be a death sentence a hundred years ago, until research in dogs identified that insulin was the influential factor, which quickly lead to treatment in humans. Both basic research (understanding how animals function), and applied research (developing drugs) are needed to make advances in the fight against diseases.
As well as this basic and applied research, any drug which has been approved for use in humans was first tested for toxicity in animals, this is a legal requirement. The testing is the result of problems identified over the years. Take thalidomide for example, this was used in humans without being tested for developmental toxicity in animals, and it turned out to cause foetal limb defects. Therefore, developmental toxicity testing in animals was introduced to check drugs had no effect on developing babies.
How is the animal work controlled and monitored to ensure it is carried safely and efficiently?
The UK has some of the tightest regulation in the world when it comes to animal testing. The UK has something called the Animal Scientific Procedures Act 1986 and also comes under new EU regulations. All work is monitored by the Home Office, every experiment is carried out under a project licence which has been reviewed by the Home Office, every individual must have a personal licence which means they’ve had accredited training, and anywhere animals are being used for scientific use must have an establishment licence. We are inspected regularly and licences can be revoked by the Home Office if necessary.
The general philosophy is that any potential benefit that each experiment could have must outweigh any potential harm to animals. The law also emphasises the rules of ‘Replacement, Reduction, Refinement’ which means we avoid using animals wherever possible, replacing them with another model, we use the least number of animals possible, reducing the number used, and we use the least distressing methods by refining the protocol. Finally the conditions in which the animals are kept and the care they receive is rigorously controlled to be of the highest standards.
Scientific animal research has been portrayed very negatively in the past, how do these portrayals differ from reality? Has the use of animals in scientific research changed a lot since?
I think the Animal Scientific Procedures Act 1986 changed things a lot, and for the better. The quality and care is what you would expect in a veterinary practice, and always has been whilst I’ve worked with animals in science. However, I think the public perception of animal use has changed. There has been a lot of negative attitudes towards scientific use of animals but that has shifted over time. Certainly, the portrayal of what happens in animal research is often not accurate at all.
One thing that’s changed over time is how open researchers are about what they do. I was always told to be careful of what I said about what I did, mainly because of the extremism which was quite prevalent then. However, society needs this research doing. We go and talk in schools and with the public, which helps to change attitudes, and we’re trying to be more open by talking about what we do.
Interview by Jodie Stephenson
Jodie is a 2nd year PhD student investigating translational biomarkers in pre-clinical models of Motor Neurone Disease. Jodie is supervised by
Dr Richard Mead and is part of the Shaw Research Group.
You can follow Jodie on Twitter @neuroruncake, LinkedIn and ResearchGate.
Tuesday, 14 July 2015
|The DNA double helix|
What is DNA, and why is it so important?
Deoxyribonucleic acid (DNA) is made up of 4 nucleotides which form rungs of a ladder-like structure called a double helix. These nucleotides can be arranged in different ways, and this is what makes up our DNA sequence. Each person’s string of DNA makes up their genome, which is unique to them, just like their fingerprint. Some diseases can be caused by some sections of this DNA being faulty, so understanding these DNA sections can be crucial in identifying disease, and could affect the treatments given to patients.
Understanding your body’s unique fingerprint: The Human Genome Project
This means that more needs to be known about the human genome. This led to the Human Genome Project (HGP) which aimed to identify the sequence of our nucleotides as well as creating maps showing the genes' location. The project began in 1990 and was funded by the US government in conjunction with many other countries, including the UK. The project was completed in 2003, with many new findings. It was found that there are around 20,500 different genes in the human genome, as well as giving information on the organisation, function and structure of our genes. That’s a lot of information!
Armed with these new findings about our healthy genes, it has allowed many genes to be identified that relate to specific diseases. Sometimes, these faults, known as mutations, have no effect on a person, but other times it can lead to disease. Some neurodegenerative diseases are known to be caused by mutations, such as: Huntington’s disease, familial motor neuron disease and familial Alzheimer’s disease, which have mutations that can be passed on through the generations, with devastating effects.
Having this knowledge is beneficial so that scientists can better understand neurodegenerative diseases, in order to develop effective drugs to target these mutations.
But has this changed the way we carry out research? Well, the answer to that question is yes! Many labs, including SITraN, are at the forefront of this work, much of it funded by the ice bucket challenge! We definitely used up a lot of ice when the whole of SITraN took part!
|The SITraN ALS/MND Ice Bucket Challenge|
Project MinE: Hope for MND Sufferers
With these advances in genome sequencing and the increasing availability of facilities to carry out sequencing at lower costs, it is now possible to apply this technique to discovering variations in a person’s DNA and see if there are any links between sufferers of neurodegenerative disease, such as motor neurone disease (MND).
This availability led to a collaboration between many countries, including the UK, USA, Switzerland and Australia, joining forces in one of the biggest human genome projects to date: Project MinE. This project, the UK arm fueled by the MND Association, aims to sequence 15,000 MND sufferer’s DNA, 1,500 of which will be sequenced right here in the UK, which is very exciting news! This information could revolutionise MND research by helping us to identify any genetic patterns that would increase the risk of someone developing this disease; it could even help to identify if any gene-environment interactions are at play, as there is evidence to suggest that lifestyle factors could have an effect on the risk of developing MND.
The results of these findings would be very beneficial and provide a huge resource to researchers that could be used in the fight against MND, with the hope of identifying which key parts of our DNA are already susceptible to MND and even if these parts could be targeted to produce a drug that could help with the symptoms and slow the progression of MND.
Mainstream genome sequencing: miracle of science?
Genome sequencing is now becoming so popular you can now pay some private companies to sequence your genome for you! This can give you lots of information, such as your susceptibility to disease. Used carefully, they could even become part of your medical records and this opens up the option of being able to tailor-make drugs that are specifically targeted at your unique genome. This may sound like a scene from a sci-fi film, but it could eventually become a reality!
So overall, the concept of mapping your own body’s fingerprint is possible and cutting edge research uses this technique to come up with novel ways of targeting and identifying markers of disease that may help us all to understand how to lead healthier lifestyles that could reduce our chance of developing these diseases either now or later on in life, as well as using this knowledge to our advantage regarding treatments if a disease has developed.
To find out more about ProjectMinE visit www.projectmine.com or the info pages on the MND Assocation website. Anyone interested into getting into this exciting field of research can check out the University of Sheffield's new
MSc in Genomic Medicine.
By Charlie Appleby-Mallinder
Charlie is an undergraduate Biochemistry student currently on a year-long placement at SITraN, supervised by Dr Paul Heath. Charlie is investigating new techniques into microdissection that can be applied to neurodegenerative research with an interest in disease at a molecular level. Charlie.firstname.lastname@example.org
Wednesday, 1 July 2015
One of the biggest discoveries in MND research in recent years has been the discovery that mutations of a gene with the rather snappy name of C9ORF72 are one of the most common causes of MND. This mutation takes the form of several thousand repeats of the sequence GGGGCC in the middle of the gene, so the sequence goes GGGGCCGGGGCCGGGGCCGGGGCCGGGGCCGGGGCC and so on for a few thousand repeats.
A gene is a code written in DNA that is the recipe for making a protein. Proteins are the building blocks and building tools for making cells, tissues, organs and bodies. To make a protein, the cell first copies the DNA gene code to RNA, then processes and prepares the RNA copy, then uses that as the template pattern for the protein.
MND researchers are spending a lot of time trying to work out how this repeat causes disease: The options are
1) the cell can’t make enough of the protein encoded by C9ORF72 because of the cumbersome GGGGCCGGGGCC (etc) repeat stuck in the middle of it.
2) The molecular tools that process RNA copies of genes get stuck to the repeat, so there aren’t enough of these tools left over to process RNA copies of other genes.
3) The cell gets confused and tries to use the GGGGCCGGGGCC repeat as a template to make rather odd proteins that shouldn’t exist and deposits them as ‘repeat inclusions’ in the cell and these are in some way toxic to the cell.
Repeating sequences in genes have been known to cause many other diseases, and the same scientific question applies: Is it poor protein production, disrupted RNA processing or trying to turn the repeat into protein that causes the problem. A very commonly repeated sequence is CAG which can cause Huntington’s disease, Kennedy’s disease (another motor neurone disease) and a number of diseases called “spinocerebellar ataxias” which result in problems of the spinal cord and cerebellum, the part of the brain at the back that aids coordination and balance. Long repeats of CAG in the gene ATXN2 cause spinocerebellar ataxia type 2.
Here’s the surprising issue: While long CAG repeats in ATXN2 cause spinocerebellar ataxia, medium-sized repeats of CAG in ATXN2 cause motor neurone disease. We looked at the brains and spinal cords from MND patients who had medium-sized repeats. Like patients with C9ORF72 they get the normal microscopic features of MND, but they don’t seem to have the repeat inclusions. Which makes us think more carefully about the other 2 options: not enough protein and disrupted RNA processing, both in ATXN2-MND and C9ORF72-MND and possibly non-inherited cases as well.
Much of the work was done by Alejandro Lorente Pons whilst studying for his MSc in Translational Neuroscience. He enjoyed this so much he stayed on with us to do a PhD in MND research – good choice Ale!
The research presented here is now published in the journal
Neuropathology and Applied Neurobiology:
J R Highley, A Lorente Pons, J Cooper-Knock, S B Wharton, P G Ince, P J Shaw, J Wood and J Kirby. Motor neurone disease/amyotrophic lateral sclerosis associated with intermediate length CAG repeat expansions in Ataxin-2 does not have 1C2-positive polyglutamine inclusions. Epub June 2015 onlinelibrary.wiley.com/doi/10.1111/nan.12254/abstract
By Dr Robin Highley, Senior Lecturer in Neuropathology
Find out more about my research on the SITraN website.
You can follow me on
and on ResearchGate.