ARTICLE IN BRIEF
Investigators have developed a new way of clearing the mutant Huntington protein that causes Huntington disease, offering a promising approach for other neurodegenerative diseases.
Researchers in Boston have helped clear in an animal model the mutated huntingtin protein, which accumulates in the neurons of people who inherit the mutation for Huntington disease (HD), causing uncontrolled movements, dementia, and emotional disorders.
Rather than disabling or destroying the protein directly, the manipulation, which involves adding an acetyl group to a specific spot on the protein, promotes autophagy, the process by which neurons break down and recycle proteins and other cell debris. The researchers report their findings in the April edition of the journal Cell.
If effective in humans, this technique might help alleviate the symptoms of other neurodegenerative diseases that involve the accumulation of proteins in neurons.
The researchers targeted the mutant huntingtin (Htt) protein for clearance by adding an acetyl group at lysine residue 444 (K444). This acetylation facilitated the movement of the protein into autophagosomes and facilitated macroautophagy in a C. elegans roundworm transgenic model of HD. When made resistant to acetylation, the mutant protein created even larger deposits in mouse brains, and was even more toxic.
The novelty of this approach lies in the way it links acetylation to degradation, according to lead author Dimitri Krainc, MD, PhD, an associate professor of neurology at Harvard Medical School and a neurologist in the Massachusetts General Hospital neurology department.
“Acetylated mutant huntingtin is preferentially recognized by a protein called p62, which directly interacts with autophagosomes,” said Dr. Krainc, who led a team of Harvard researchers from the Massachusetts General Institute for Neurodegenerative Disease (MIND). “Therefore we think that p62 delivers' acetylated huntingtin to autophagosomes for degradation.”
He wasn't sure, however, why the acetylation process affected only mutant forms of the huntingtin protein. “We don't fully understand that,” he said. “It seems that accumulation of mutant Htt promotes its own acetylation. Normal Htt does not accumulate. It is also possible that acetylated normal Htt gets degraded faster and is therefore not detectable. We are currently investigating this question.”
A NOVEL APPROACH
Stimulating autophagy to rid neurons of unwanted proteins provides an exciting new way of dealing with certain neurodegenerative diseases, according to Ira Shoulson, MD, Louis C. Lasagna Professor of Experimental Therapeutics and Professor of Neurology, Pharmacology and Medicine at the University of Rochester School of Medicine. “I'd say it's not ready for prime time, but it's a whole new area of great therapeutic interest for Huntington disease and, I suspect, for other neurodegenerative disorders,” Dr. Shoulson said.
“If this works in Huntington disease, I see no reason why it wouldn't help in Parkinson disease, Alzheimer disease, and so on. What they've shown is that one can actually stimulate autophagy through acetylation, and that's a sensible, thoughtful approach, one that will lead to a variety of drugs to stimulate autophagy.”
MORE ON AUTOPHAGY
A group in England led by David C. Rubinsztein, MB, PhD, professor of molecular neurogenetics at the University of Cambridge, has stimulated autophagy with rapamycin, an immunosuppressant drug widely used to prevent rejection of transplanted kidneys. The treatment has successfully removed mutant huntingtin protein from neurons in animal models of the disease, including flies, zebrafish and mice. In a series of papers, Dr. Rubinsztein and his colleagues have also characterized a number of alternative drugs, including lithium, valproate and carbenazepine, which induce autophagy via a pathway that is independent of the target of rapamycin. These drugs also protect against mutant huntingtin toxicity in animal models of the disease. Furthermore, it is possible to obtain stimulation of autophagy and increased clearance of mutant huntingtin by using rapamycin along with drugs acting independently of the target of rapamycin, according to Dr. Rubinsztein.
But stimulating autophagy in the brain can have unforeseen and unwanted consequences according to Philipp A. Jaeger, co-author of a paper in the April edition of Molecular Neurodegeneration on the role of autophagy in neurodegeneration.
“Autophagy is a process that affects a lot of things, so when you try to manipulate it, you can cause a lot of changes you cannot control,” said Jaeger, who coauthored the paper with his graduate advisor, Tony Wyss-Coray, PhD, associate professor of neurology and neurological sciences at Stanford School of Medicine. “The Cell paper suggests that targeted acetylation of a single amino acid on a protein could be a way to cause the specific and effective degradation of that protein by autophagy. One would not have to manipulate autophagy as a whole, which is much harder to control.”
Dr. Rubinsztein pointed out, however, that drugs that induce autophagy do not appear to cause obvious autophagy-related side-effects. “For instance, rapamycin induces autophagy and has been used for decades as a long-term immunosuppressant in patients who have kidney transplants, and its side-effects appear to be unconnected with autophagy,” Dr. Rubinsztein said. “I would say that the general view in the autophagy field is that autophagy is considered to be a cytoprotective process in the main.”
Although the Rubinsztein group has successfully rescued neurons from the neurotoxic effects of huntingtin protein, and has also shown that tau protein can be degraded by autophagy, Jaeger still worries that stimulating autophagy in general “might lead to other changes that are hard to foresee.”
For example, while autophagy clearly helps neurons remain healthy, it also appears to cause harm in the wake of traumatic brain injury.
“We know that activating autophagy helps to clear neurotoxic proteins, but in other instances, such as following acute injury, autophagy seems to contribute to a cell death pathway,” Jaeger said. “It actually speeds up the death of neurons and should therefore be inhibited.”
Dr. Krainc's approach, in contrast, stimulates the removal of a specific toxic protein without stimulating autophagy itself. This suggests that unwanted proteins might be tagged for destruction by other processes in addition to acetylation.
“For example, in Alzheimer disease tau is hyperphosphorylated,” Jaeger said. “Phosphorylation is somewhat similar to acetylation — it's the modification of an amino acid through the addition of a phosphorous group. It will be very interesting to see if tau is being degraded by autophagy, and if phosphorylation events affect that degradation the way acetylation affects huntingtin degradation.”
ANOTHER WAY TO CLEAR MUTANT HTT
While promoting autophagy of mutant Htt gets rid of the protein after it is formed, researchers at the University of Texas are trying to block the production of Htt by targeting the “hairpin” formed by CAG repeats on the mutant gene.
The gene that produces normal Htt contains between 12 and 39 repeats of CAG. People with Huntington disease carry a gene that contains more than 45 CAG repeats, which produces a mutant form of Htt that accumulates in neurons, causing dysfunction and death.
Since people with HD carry one normal and one mutant allele for Htt, the long CAG repeat in the mutant form might provide a target for oligomers that can disrupt its transcription, reasoned David C. Corey, PhD, professor of pharmacology and biochemistry at the Medical Center, and lead author of a paper published online May 3 in advance of the print Nature Biotechnology.
For about 10 years Dr. Corey has been working with peptide nucleic acids that function as antisense molecules capable of modifying the expression of genes. The Hereditary Disease Foundation, a nonprofit dedicated to finding cures for genetic diseases, asked him if he would be interested in trying to disrupt the expression of the gene that causes HD.
“I had the idea that the CAG hairpin will be short in the wild type gene and long in the mutant gene, so by definition the energies of the short wild type and the long mutant would be different,” Dr. Corey said. “And because they're different they might have a different ability to bind to an oligonucleotide. We had no idea if that would be true, but we thought we'd try it, and it turns out it works.”
Dr. Corey found that a locked nucleic acid blocked the expression of mutant Htt but did not affect the expression of other genes containing a smaller number of CAG repeats.
“Locked nucleic acid is a molecule that's already being tested, so it's not such a stretch to think a company might be interested in developing it as a treatment,” Dr. Corey said. “Our role now is to explore some obvious modifications and see if we can improve the compounds we have. The next step would be to take the best of those compounds and try it out in an animal model and see what happens.”•
* Acetylation: The addition to a molecule of an acetyl group consisting of oxygen, hydrogen, and carbon atoms.
* Phagosome: A vacuole that engulfs and digests foreign matter brought into a cell.
* Autophagosome: A phagosome that digests material in the cell's own cytoplasm, setting in motion the process of autophagy.
* Autophagy: The process by which cells break down and recycle their own proteins by passing them through lysosomes within the cell.
* Macroautophagy: The process by which proteins in a cell are sequestered by an autophagosome, which fuses with a lysosome and degrades the proteins.
* Microautophagy: The process by which the lysosome directly ingests and degrades unwanted proteins and other material.
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