New molecule can reverse the Huntington’s disease mutation in lab models

Written by Dr. Michael Flower Edited by Dr. Rachel Harding

Editor’s Note: This article was initially published by HDBuzz on March 13, 2020. They have graciously allowed us to build on their work and add a section on how this research may be relevant to ataxia. This additional writing was done by Celeste Suart and edited by David Bushart.

A collaborative team of scientists from Canada and Japan have identified a small molecule which can change the CAG-repeat length in different lab models of Huntington’s disease.

CAG repeats are unstable

Huntington’s disease is caused by a stretch of C, A and G chemical letters in the Huntingtin gene, which are repeated over and over again until the number of repeats passes a critical limit; at least 36 CAG-repeats are needed to result in HD.

In fact, these repeats can be unstable, and carry on getting bigger throughout HD patients’ lives, but the rate of change of the repeat varies in different tissues of the body.

In the blood, the CAG repeat is quite stable, so an HD genetic blood test result remains reliable. But the CAG repeat can expand particularly fast in some deep structures of the brain that are involved in movement, where they can grow to over 1000 CAG repeats. Scientists think that there could be a correlation between repeat expansion and brain cell degeneration, which might explain why certain brain structures are more vulnerable in HD.

a print out of genetic information show as a list of A,T, C, and G letters
The CAG repeat of the huntingtin gene sequence can be changed to include more and more repeats, in a process called repeat expansion. This can also happens in some ataxia related genes. Image credit: “Gattaca?” by IRGlover is licensed under CC BY-NC 2.0

But why?

This raises the question, what is it that’s causing the CAG repeat to get bigger? It seems to be something to do with DNA repair.

We’re all exposed continually to an onslaught of DNA damage every day, from sunlight and passive smoking, to ageing and what we eat. Over millions of years, we’ve evolved a complex web of DNA repair systems to rapidly repair damage done to our genomes before it can kill our cells or cause cancer. Like all cellular machines, that DNA repair machinery is made by following instructions in certain genes. In effect, our DNA contains the instructions for repairing itself, which is quite trippy but also fairly cool.

What is it that’s causing the CAG repeat to get bigger? 

We’ve known for several years that certain mouse models of HD have less efficient systems to repair their DNA, and those mice have more stable CAG repeats. What’s more, deleting certain DNA repair genes altogether can prevent repeat expansion entirely.

But hang on, isn’t our DNA repair system meant to protect against mutations like these?? Well normally, yes. However, it appears a specific DNA repair system, called mismatch repair, sees the CAG repeat in the huntingtin gene as an error, and tries to repair it, but does a shoddy job and introduces extra repeats.

Why does this matter?

There’s been an explosion of interest in this field recently, largely because huge genetic studies in HD patients have found that several DNA repair genes can affect the age HD symptoms start and the speed at which they progress. One hypothesis to explain these findings is that slowing down repeat expansion slows down the disease. What if we could make a drug that stops, or even reverses repeat expansion? Maybe we could slow down or even prevent HD.

So what’s new?

Chris Pearson’s group in Toronto have developed a compound called naphthyridine-azaquinolone, which we’ll just refer to more easily as ‘NA’, which binds CAG repeats and could prevent repeat expansion.

Using cells from HD patients in a tissue dish, NA was shown to successfully slow, and possibly even lead to a small reduction in CAG repeat length. Pearson showed that blocking transcription, the process in which genes are used as templates to make proteins, prevents repeat expansion. This suggests that during transcription, the huntingtin repeat might be bent into an abnormal shape, which mismatch repair machinery in the cell recognises and then tries to repair. However, precisely how NA works in this process remains unclear.

Pearson’s team injected NA into one side of the brain of an HD mouse model. They targeted the striatum, a region known to show lots of CAG expansion. Compared to the untreated side, NA prevented expansion and even caused some shrinkage of the repeat number.

Next, they showed NA reduced the build-up of clumps of toxic huntingtin protein in the mice’s cells. It is not clear yet whether the treated mice have improved symptoms or increased lifespan. This will be important for scientists to work out before deciding whether preventing repeat expansion has potential as a therapy for people.

Could NA also be used in ataxia?

As part of their experiments, Pearson’s team tested to see if NA would affect proteins with stable CAG/CTG repeats that don’t tend to expand. This included two ataxia related genes, ATXN8 (mutated in SCA8) and TBP (mutated in SCA17). When cells were treated with NA, it didn’t have any impact on the repeat length of ATXN8 and TBP. Both stayed the same length they were to begin. That means NA wouldn’t be useful for these forms of ataxia.

The next question is if NA could be used for CAG repeat ataxias that also have unstable repeats. Genes involved in SCA1, SCA2, SCA3, and SCA7 have all been shown to sometimes grow in length like the huntingtin gene. Pearson’s team didn’t test how NA impacts these genes directly in the paper, so we don’t know for certain.

There is one big difference between these ataxias and HD: The areas of the brain where there is repeat expansion in huntingtin are also the places degeneration happens. But in SCAs, these expansions don’t always line up with where there is degeneration. In the cerebellum, where the most degeneration occurs in ataxia, CAG repeats don’t expand very much. In fact, some researchers have even found slightly shorter repeats in the ATXN1 gene (mutated in SCA1), the ATXN2 gene (mutated in SCA2), and the ATXN3 gene (mutated in SCA3) in the cerebellar cortex than in other parts of the brain and body.

We don’t know why repeats seem to more stable in the cerebellum. But since ataxia is a disease of the cerebellum, the finding that we don’t see repeat expansion in this section of the brain has caused some ataxia researchers to move away from researching this to looking at other hypotheses.  But we do see degeneration caused by ataxia in other areas of the brain, like the brainstem and midbrain. It might be useful to revisit NA once we find out more about how these regions play a role in ataxia.

As work continues on researching NA and its effect on repeat expansion in Huntington’s disease, Pearson’s team will likely do preliminary testing to see if there are any effects in related ataxias. When that happens, we will know for certain if NA can prevent the expansion of CAG repeats and slow down symptom progression in spinocerebellar ataxias.

What’s the catch?

NA was shown to successfully slow, and possibly even lead to a small reduction in CAG repeat length 

A huge obstacle to making new drugs is getting them into the cells that most need them; in the case of HD, that means throughout deep regions of the brain. NA is able to freely enter different cells once in the brain, but this current version of the molecule has not yet been shown to cross the blood-brain-barrier. Scientists might need to modify and improve the NA molecule to avoid needing to be directly injected into the brain.

Fiddling around with DNA repair, one of our body’s major defence systems, could be dangerous, and there’s the potential for major side effects like cancer. Pearson showed that NA didn’t affect the core function of mismatch repair, which is to remove DNA bases when they get put in the wrong place. The researchers carefully analyzed the rate of mutations across the whole genome, and there was no detectable increase in the rate at which they were found, compared to controls when they were treated with NA.

It is possible to imagine treating HD patients at an early age, before they develop any symptoms; this might stabilise the CAG repeat and could prevent or at least delay the onset. CAG repeat shrinkage in their sperm or eggs could even mean they wouldn’t pass the disease on to their children.

However, for NA there is still a lot of work to do. For starters, we would need to show that preventing CAG expansion slows down the disease, we would then need to come up with a way to get NA into the deep regions of the brain, and finally we would need to be sure it is safe with limited side-effects. Early treatment could also mean being exposed to risks like cancer for even longer, so there’s clearly a lot to be worked out.

In summary, NA is an exciting research compound, but there is still a long road ahead before something like it might be a drug that could be taken by people to prevent or treat Huntington’s disease.

Key Terms

CAG repeat: A stretch of DNA that was the sequence CAG repeated many times. Everyone has repeating CAG tracts in some genes, but once they are over a certain length they can lead to disease, like Huntington’s Disease. Some ataxias are also caused by this type of mutation, including SCA1, SCA2, SCA3, SCA6, SCA7 and SCA17.

Genome: A collection of all the genes that have the instructions needed to make an organism or person

Transcription: The process in which the genetic information from DNA is copied into RNA, another medium for genetic information. To learn more about transcription, see our Snapshot on What is RNA.

Conflict of Interest Statement

Dr. Michael Flower, the original writer of this article, declares no conflicts. Dr. Rachel Harding, the original editor of this article, is a collaborator of the Pearson laboratory but has no connection to this published study.

Celeste Suart, who wrote the section on ataxia related information, is from a laboratory which collaborates with the Pearson laboratory but has no connection to this published study. David Bushart has no conflict of interest to declare.

Citation of Article Reviewed

Nakamori, M., Panigrahi, G.B., Lanni, S. et al. A slipped-CAG DNA-binding small molecule induces trinucleotide-repeat contractions in vivo. Nature Genetics 52, 146–159 (2020). doi: 10.1038/s41588-019-0575-8.