Surprising way genetic mutations cause Huntington's disease changes our understanding of Huntington's disease

Scientists at MIT and Harvard's Broad Institute, Harvard Medical School, and McLean Hospital have discovered a surprising mechanism by which the genetic mutation known to cause Huntington's disease causes brain cell death. The findings change our understanding of the deadly neurodegenerative disease and suggest ways to potentially slow or prevent it.

Researchers have known for 30 years that Huntington's disease is caused by genetic mutations in the huntingtin (HTT) gene, but they did not know how the mutations were caused. brain cell death.

Research published in cell It turns out that the inherited mutations themselves do not harm the cells. Rather, this mutation is harmless for decades, but slowly transforms into a highly toxic form that quickly kills cells.

The Huntington mutation involves a stretch of DNA in the HTT gene in which the three-letter sequence “CAG” of DNA is repeated at least 40 times. In comparison, people without the disease inherit between 15 and 35 repeats. The researchers found that DNA tracts with more than 40 CAG repeats grew to lengths of several hundred repeats.

This type of “somatic cell expansion” occurs only in certain types of brain cells that later die from Huntington's disease. Only when a cell's DNA expansion reaches a threshold number of CAGs (approximately 150) does the cell become diseased and subsequently die. The cumulative death of many of these cells causes the symptoms of Huntington's disease.

The study offers a potential explanation as to why Huntington's disease drug candidates aimed at reducing HTT protein expression have struggled in clinical trials. Few cells have a toxic version of the protein at any given time, so treatments may not be working properly. therapeutic effect in most cells.

This research also improves alternative treatment strategies. Stopping or slowing the expansion of the CAG repeats in the HTT gene could delay toxicity in a much larger number of cells and potentially delay or prevent disease onset.

“These experiments have changed the way we think about how Huntington's disease develops,” said Steve McCarroll, a geneticist and neuroscientist and co-senior author of the study. said.

McCarroll is a member of the institute, director of genomic neurobiology at the Stanley Psychiatric Research Center at the Broad, the Dorothy and Milton Fryer Professor of Biomedical Sciences and Genetics at Harvard Medical School, and the Howard Hughes Medical Research Institute. He is also a researcher at the institute.

“This is a completely different way of thinking about how mutations cause disease, and we think it can be applied to DNA repeat disorders other than Huntington's disease.”

“The point of all of our work is to alleviate the suffering caused by disease,” said co-author Sabina, associate professor of psychiatry at McLean Hospital, a member of Harvard Medical School and the Massachusetts General Brigham Health System. Beretta said. .

She is also the director of the Harvard Brain Tissue Resource Center (HBTRC), an NIH neurobiobank center at McLean Hospital. “This study and the research it informs is impactful and could make a big difference in reducing suffering in the short term.”

Bob Handsaker, a staff scientist in McCarroll's group, Seva Kashin, a senior principal software engineer, and Nora Reed, a former researcher, are all co-lead authors on the study.

unanswered questions

Huntington's disease destroys a population of cells called striatal projection neurons located in the striatum, a deep brain structure responsible for movement, many cognitive functions, and motivation.

When these cells die in large numbers, patients develop involuntary movements in the arms, legs, and face, and many also develop cognitive impairment. These symptoms usually begin in middle age and progress over 10 to 20 years, resulting in more severe cognitive impairment and difficulty moving and swallowing.

In 1993 researchers discovered The disease is caused by an expansion of the CAG within the HTT gene. Most people who inherit a version of the gene with 15 to 35 consecutive CAGs do not develop Huntington's disease, but those who inherit a version with 40 or more consecutive CAGs are most likely to develop Huntington's disease later in life. The disease develops in 2018.

The longer the period of recurrence, the younger the symptoms tend to first appear. It has also been shown that the canals of repeated CAGs expand over time and have varying lengths in different tissues.

But fundamental biological questions lingered. “How are HTT mutations toxic?” Why does the HTT protein, which is present in nearly every cell in the body, kill only some brain cells and not others? And Why do patients who are born with a mutation and express the protein throughout their lives only develop symptoms in middle age, after appearing healthy for decades?

Repeat expansion

To answer these questions, the research team turned to droplet single-cell RNA sequencing, developed a decade ago by the McCarroll lab.drop sequence) This allows researchers to analyze gene expression in thousands of single cells.

In an effort to understand the direct biological effects of CAG repeat length, researchers conducted single-cell RNA sequencing to be able to determine not only gene expression and the identity of single cells, but also the length of DNA repeat tracts within each cell. was applied.

“These repeats are known to expand within neurons,” Kassin says. “But being able to take specific cells and measure both the CAG length and the transcriptional profile is a very important foundation that allows for very powerful analyses.”

The researchers studied brain tissue provided by 53 people with Huntington's disease and 50 patients with Huntington's disease, collected and archived by the HBTRC.

They analyzed more than 500,000 single cells and found that most cell types in people with the disease have essentially the same CAG repeats as the ones they inherited. However, striatal projection neurons (the primary striatal cells that die in disease) had significantly expanded CAG repeat pathways.

Most previous studies of human brain tissue focused on CAG repeat regions with fewer than 100 repeats, but new research shows that some neurons have as many as 800 CAGs. discovery It was created 20 years ago by Peggy Shelbourne at the University of Glasgow.

Most surprisingly, the researchers found that expanding the DNA repeats from 40 to 150 CAGs had no apparent effect on neuron health. However, neurons with more than 150 CAG of repeats exhibited highly skewed gene expression, lost expression of important genes, and subsequently died.

McCarroll's team also used computer modeling of data to estimate the speed and timing of CAG repeat expansion in striatal projection neurons.

They found that the CAG repeat region initially grows slowly, expanding less than once a year during the first 20 years of life. However, once the cell repeat tract reaches approximately 80 CAG, usually after several decades, its rate of expansion accelerates dramatically, expanding to 150 CAG in just a few more years.

The cells then die after a few months. This means that a neuron spends more than 95% of its life with a harmless HTT gene. Furthermore, because CAG repeat tracts in different cells cross this toxicity threshold at different times, cells as a population disappear slowly over a long period of time, starting approximately 20 years before symptoms appear, and disappear more quickly once symptoms begin.

“While much was known about Huntington's disease before we began this study, there were gaps and inconsistencies in our overall understanding,” Handsaker said. “We were able to piece together the complete trajectory of pathology that develops over decades in individual neurons, which could provide different time points at which we can intervene therapeutically.”

Analysis of brain tissue donated by Huntington's disease patients was critical to this study. “I'm grateful to the family for choosing to do something so difficult,” Beretta said. “This would not have been possible without the altruistic spirit of the many brain donors who left behind a legacy of knowledge that will endure and benefit many others.”

Treatment possibilities

Rather than targeting the HTT protein, McCarroll's team believes that a complementary or potentially better treatment could be to slow or stop the expansion of DNA repeats, thereby slowing or preventing disease progression. It suggests that it may be possible to do so.

Previous genetic studies of Huntington's disease include: the study Vanessa Wheeler and Ricardo Mouro Pinto of Massachusetts General Hospital suggest ways to potentially slow the spread.

Studies have shown that cellular proteins involved in DNA maintenance and repair can compromise the stability of DNA repeat tracts. For example, the MSH3 protein normally helps cells monitor potential mutations in their DNA, but loops in the DNA formed by extra CAGs can disrupt this protein and cause expanded CAG repeats. There is a gender.

An international team of human geneticists Found Dr. McCarroll said common genetic variations in the genes that code for these DNA repair proteins may cause Huntington's disease patients to develop symptoms earlier or later. This discovery directly prompted the research team to focus on developing a method to measure CAG repeats in single cells.

He added that slowing down certain DNA maintenance processes with molecular therapies could allow other, less error-prone DNA repair mechanisms to resolve these loops, slowing the expansion of DNA repeats. .

Meanwhile, researchers are working to understand how DNA repeat tracts larger than 150 CAG lead to neuronal damage and death, and why repeats are more expanded in some types of neurons than in others. I'm here. They also combined DNA repeat profiling with similar single-cell RNA sequencing to understand the association between DNA repeat expansion and cellular changes in other genetic diseases involving late-onset DNA repeats in patients. I am using it.

More than 50 human brain diseases, including fragile X syndrome and myotonic dystrophy, are caused by expansions of DNA repeats in various genes.

“It will take a lot of scientific research by a lot of people to develop treatments that slow the expansion of DNA repeats,” McCarroll said. “But we hope that understanding this as a central process that drives disease will lead to deeper focus and new options.”