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Investigators identified a novel disease pathway — RAS-MAPK-MSK1 — for spinocerebellar ataxia type 1 by looking for common genetic hits in multiple species. The technique may apply to research on other neurodegenerative disorders, experts said.
A team of investigators at Baylor College of Medicine and the University of Minnesota has identified a new disease pathway for spinocerebellar ataxia type 1 (SCA1) by screening for common genetic hits for the disorder across multiple species. The unique approach, called complementary forward genetic screening, could be used to find pathways at work in other neurodegenerative diseases and lead to novel therapeutics, experts said.
Huda Y. Zoghbi, MD, and her colleagues at Baylor College of Medicine and the University of Minnesota had previously identified the genetic mutation for SCA1, showing the disease is caused by expansion of a polyglutamine tract in ataxin 1 (ATXN1). Too much protein, abnormal or even wild type, causes the protein to misfold and pile up until it reaches a critical mass and becomes toxic to the cell. Phosphorylation of a single amino acid is critical in this process.
But even with more than a decade of scientific exploration, their tools failed to identify the pathway responsible for modulating ATXN1 protein levels and disease toxicity. Genetic screens of several model systems — a SCA1 drosophila model and human cell models of the disease — offered them a window into shared genes that target the levels of glutamine-expanded human ATXN1.
They focused their initial search on genes that make over 600 human kinases because they knew that kinases are involved in phosphorylation and are good targets for drug development. It was a risky study with equal odds of working or not, but the idea paid off.
They identified 10 genes that all link back to the RAS-MAPK-MSK1 pathway, and were able to show they play an important role in modulating ATXN1 protein levels. Down-regulation of several components of the RAS-MAPK-MSK1 pathway, they found, decreased ATXN1 levels and suppressed neurodegeneration in drosophila and mice. Further experiments also proved that drugs that targeted the kinases in this pathway reduced ATXN1 levels. The findings were reported in the May 29 online edition of Nature.
The findings could open a door to new therapeutic “entry points” for SCA1 and other neurodegenerative diseases, said Howard Hughes Medical Investigator Dr. Zoghbi, a professor of pediatrics, molecular and human genetics, neurology, and neuroscience at Baylor College of Medicine, and director of the Jan and Dan Duncan Neurological Research Institute at Texas Children's Hospital.
The RAS-MAPK-MSK1 pathway is induced in cells when they undergo stress. The pathway is well studied in fields of inflammation, immunity, and cancer. The investigators are now looking beyond kinases to test other genes. The hope is that identifying other pathways involved early on in the disease process would allow scientists to develop drugs that tweak the pathways to nudge protein levels up or down in small ways rather than take a therapeutic sledgehammer to one pathway.
“This approach may be less toxic,” said Dr. Zoghbi. “We can use this strategy to modulate any protein,” she added. “It takes you further upstream to the problem.”
Like many neurodegenerative conditions, SCA1 takes decades before symptoms develop. Clinical signs develop in the third decade. Patients have trouble walking and talking, bodily functions that had seemed normal are no longer, and life is cut short.
Earlier findings from the investigators suggested that lowering ATXN1 levels by 20 to 50 percent was enough to reduce many of the clinical symptoms in animal models of the disease. This finding was exciting for SCA1 but also held out hope for other neurological conditions caused by abnormal protein misfolding and accumulation that leads to cell death.
Even slight modulations of the RAS-MAPK-MSK1 pathway were enough to reduce the levels of defective ATXN1, said Harry T. Orr, PhD, a long-time collaborator of Dr. Zoghbi who is director of the University of Minnesota Institute for Translational Neuroscience. “Perhaps, if you decrease the levels of the protein, you will decrease the severity of the disease.” Collaborators of the study include Baylor's Juan Botas, PhD, and Thomas Westbrook, PhD; post-doctoral fellow Jeehye Park, PhD, who works in Dr. Zoghbi's lab; and Ismail Al-Ramahi, PhD, a postdoctoral fellow in the Botas lab.
The collaboration was key. “Harry and I had studied the disease and we had animal models. Juan Botas, a professor of molecular and human genetics at Baylor College of Medicine had a fruit fly model, and Dr. Westbrook had a nice technology that enabled us to monitor ataxin-1 levels,” Dr. Zoghbi explained. Dr. Park and her colleagues carried out the screen in human cell lines and Dr. Al-Ramahi and his colleagues carried out the screen in the fruit flies.
Once the new pathway was identified they focused on the MSK1 protein in this pathway and found that decreases in the level of the protein in mice with SCA1 resulted in reduced levels of ATXN1. Most important, the animals improved, said Dr. Zoghbi. “This final experiment proved that reducing levels of the protein could stave off the disease.”
“Now that we know that it works with ataxin 1, we can conduct similar studies with many proteins whose levels drive neurodegeneration in sporadic and inherited diseases such as Alzheimer's, Parkinson's, Huntington's and other neurological disorders,” said Dr. Zoghbi.
“This is getting us really close, not only for SCA1, but I think it's going to be a guidepost for work on a lot of other neurodegenerative diseases,” Dr. Orr said.
The research was funded through a new program grant from the Howard Hughes Medical Institute's Collaborative Innovation Awards program. It was intended to allow teams of investigators from many fields to come together to tackle high-risk projects.
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Kenneth H. Fischbeck, MD, chief of the neurogenetics branch at the National Institute of Neurological Disorders and Stroke (NINDS), said that the novelty in this study is that the investigators used two different approaches: RNA interference (RNAi) to knock down genes in culture, and then they looked in the fly model of SCA1 to identify genes being expressed. They identified 45 of 636 genes for kinases in human cell culture and 51 of 337 genes in the flies. They focused on the 10 genes that overlapped in human cell culture and in the flies.
“It is an elegant way of using two model systems,” said Dr. Fischbeck. “It makes a strong case for this particular set of genes in patients with SCA1.”
He said that the next step is to see whether this pathway is involved in human forms of the disease. “The same kind of approach can work for any disease where we know the mechanism. It is a model that the rest of us can follow.”
David M. Holtzman, MD, the Andrew B. and Gretchen P. Jones professor and chairman of neurology, professor of developmental biology, and associate director of the Alzheimer's Disease Research Center at Washington University in St. Louis, agreed that the research strategy could bear fruit for other neurodegenerative disorders.
“It is clear that decreasing the levels of certain proteins that aggregate in different neurodegenerative diseases is very likely to be a good future strategy for developing new preventions/treatments for neurodegenerative diseases such as Alzheimer's, Parkinson's, Huntington's, ALS, SCA1, etc.,” said Dr. Holtzman, who was not involved with the study. “The Zoghbi lab used an interesting approach in a proof-of-concept fashion to figure out what genes/pathways can result in a decrease in SCA1 levels. By using an unbiased approach, they found a pathway (RAS-MAP kinase) that regulated SCA1 levels and thus SCA1-related pathology.
“The implication is that a similar approach could also be utilized to identify this or other pathways that might be involved in regulating levels of other key proteins in neurodegeneration such as APP [amyloid precursor protein], tau, synuclein, etc. I think this approach may in fact be a very good way to identify novel pathways to regulate the level of these proteins. If it works, one might be able to then develop novel therapeutics depending on the target pathways one identifies.”