Lewy Bodies Diverse in Structure and Some Fibrils Can Migrate, Study Reports

alpha-synuclein study

A detailed analysis of the structure of alpha-synuclein clumps suggests that Parkinson’s is a systemic disease, whose characteristic protein aggregates can move about inside the brain and migrate beyond the central nervous system, according to a new research.

This finding may help in better understanding why Parkinson’s patients experience symptoms other than the disease’s characteristic motor problems.

The study, “Parkinson’s disease is a type of amyloidosis featuring accumulation of amyloid fibrils of α-synuclein,” was published in the journal Proceedings of the National Academy of Sciences.

A hallmark feature of Parkinson’s is the accumulation of small and complex structures called Lewy bodies, which are mainly composed of the alpha-synuclein protein in nerve cells of the brain. Recent work has shown these aggregates can travel across cells of connected brain regions. But little is known about how this migration is regulated, and scientists are still working to more fully understand the structure of proteins in Lewy bodies.

A team from Osaka University, in Japan, used a technique called microbeam X-ray diffraction to gather information in greater detail about the structure of alpha-synuclein clumps.

Using this technique, researchers can detail the complex 3D structure of protein aggregates based on the diffraction pattern they produce when crossing a beam of X-rays. (Diffraction patterns here refer to the bending of X-ray waves as they pass an object.)

The team first tested the sensitivity of their approach using senile plaques from mice in a model of Alzheimer’s disease. These plaques are also composed of complex protein clump, but consist of the beta-amyloid protein.  The researchers then used the same method to analyze thin brain sections taken from three Parkinson’s patients who died between the ages of 75 and 83.

Tests confirmed that protein clumps from patients mainly consisted of alpha-synuclein, by using an antibody specific for detecting this protein. Next, the researchers saw different X-ray scattering patterns in mice and patient tissue samples, confirming they consisted of different proteins.

They also found that different patient samples had slightly different scattering patterns, suggesting diverse clump structures. Importantly, some structures seen in patients’ alpha-synuclein aggregates were similar to structures previously reported in mice studies, which found fragments of these protein fibrils (called cross-beta, or cross-β, structures) could propagate — migrate — throughout the body.

“Our study is the first to find that aggregates in Parkinson’s disease brains also have this cross-β structure,” Hideki Mochizuki, MD, PhD, the study’s senior author, said in a news release.

Inconsistent findings across patient samples might indicate “the different maturity stages of Lewy bodies,” said Katsuya Araki, MD, PhD, and the study’s first author.

Importantly, rather than supporting Parkinson’s as a disease localized in the brain, “our finding supports the concept that [Parkinson’s] is a type of amyloidosis, a disease featuring the accumulation and propagation of amyloid fibrils” of alpha-synuclein, they wrote.

This appears to be in line with both the non-motor symptoms experienced by Parkinson’s patients before difficulties with movement are manifest, and with the presence of alpha-synuclein deposits found in peripheral nerves of the heart and the gut.

“This has obvious implications in the diagnosis of Parkinson’s disease, and could also have therapeutic implications in the long run,” Araki said.

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Engineered Protein Binds to Alpha-synuclein to Prevent Toxic Clumping, Study Reports

alpha-synuclein, AS69

An engineered protein known as AS69 is able to bind to individual units of alpha-synuclein to prevent them from clumping, and — in a fly model of Parkinson’s disease — its use led to preserved motor function, a study reports.

The study, “An engineered monomer binding-protein for α-synuclein efficiently inhibits the proliferation of amyloid fibrils,” appeared in the journal eLIFE.

Cellular aggregates, or clumps, of the protein alpha-synuclein are an established hallmark of Parkinson’s and focus of research into treatments. “The link between [alpha]-synuclein aggregation and PD [Parkinson’s] has been known for two decades … however, translation of this scientific discovery into a therapy has proven challenging,” the scientists wrote.

A team of international researchers now engineered a binding protein, known as beta-wrapin AS69, that can bind with high affinity to monomers (single units) of alpha-synuclein and induce a specific conformational or structural change called a beta hairpin.

The alpha-synuclein region that adopts this structure is essential for its clumping, as indicated by the presence of a cluster of disease-related mutation sites. Upon binding to this region, AS69 stops alpha-synuclein from aggregating into amyloid fibrils.

To better determine the potential of AS69 as a therapy, the scientists also tested it in cellular and animal models.

In vitro, the team confirmed that AS69 specifically worked to lower alpha-synuclein aggregation and not its amount, testing both with wild-type (normal) protein and a variant (A53T) previously linked to familial Parkinson’s and to quicker protein clumping. (In vitro refers to experiments in lab dishes; in vivo experiments are those within a living organism, including animal models.)

In fruit flies with A53T alpha-synuclein in their brain nerve cells, AS69 was then seen to preserve the flies’ ability to climb (reflecting motor function), which was associated with fewer alpha-synuclein aggregates. This climbing ability progressively declined in the absence of AS69.

Protein aggregation is a complex process involving multiple microscopic steps. It starts with an event called primary nucleation, in which misfolded (altered shape) proteins clump together to form fibrils, which then elongate. This first step proceeds slowly, potentially taking up to several decades.

A later event is called secondary nucleation. Here, aggregation speeds up and exponential growth occurs, with existing clumps promoting the formation of new ones. This faster phase is associated with evident disease, and a potential for rapid progression.

When the team investigated specific steps of alpha-synuclein protein clumping, it found that fibril elongation was suppressed by AS69 in a concentration-dependent manner. Both fibrils and AS69 competed for the single units of alpha-synuclein, but while the interaction with AS69 occurred within seconds, binding to fibrils took minutes to hours.

In contrast, the interaction of free AS69 with fibrils was weak, if it existed at all.

AS69 was also found to interfere with lipid (fat)-induce alpha-synuclein aggregation, and was a more efficient inhibitor of the amplification of alpha-synuclein amyloid fibrils than beta-synuclein — a protein known to suppress alpha-synuclein clumping.  Importantly, by binding to alpha-synuclein, AS69 also prevented secondary nucleation.

Based on these results, the team proposed that the complex of AS69 with alpha-synuclein incorporates into a fibril precursor, and prevents this precursor from undergoing the structural changes needed for further aggregation.

“An inhibitor functioning according to this dual mode, that is being active both as a free molecule and as a complex with (…) [alpha]-synuclein, is expected to efficiently reduce [alpha]-synuclein aggregation in vivo,” the researchers concluded.

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New Compound Can Prevent Formation of Toxic Alpha-Synuclein Aggregates, Animal Study Finds

A newly identified small molecule compound, called SynuClean-D, can prevent the formation of toxic alpha-synuclein aggregates, disrupt the propagation of amyloid fibrils, and rescue nerve cells from alpha-synuclein-induced death in an animal model of Parkinson’s disease, a study shows.
Given this particular activity, SynuClean-D may be a promising candidate for the development of new Parkinson’s therapies.
The study, “Small molecule inhibits α-synuclein aggregation, disrupts amyloid fibrils, and prevents degeneration of dopaminergic neurons,” was published in the Proceedings of the National Academy of Sciences.
The process of amyloid aggregation that characterizes several neurodegenerative disorders can be simulated in the laboratory. This strategy can be used to identify and test potential therapies. However, fibril growth occurs very slowly in a highly variable manner, which can make targeted-compound screening a difficult process.
A team led by researchers at the Universitat Autònoma de Barcelona implemented a robust high-throughput assay that allows the screening of large chemical libraries in search of inhibitors that can specifically prevent alpha-synuclein aggregation.
This assay detects a compound called thioflavin-T, or Th-T, which binds to amyloid proteins and gives a strong fluorescent signal that works as a readout of amyloid formation. The team used this new method to screen about 14,400 chemical compounds in the HitFinder Collection from Maybridge, a platform with a wide portfolio of chemistry products and services specifically tailored to the drug discovery and biotechnology sectors.
Through this analysis, researchers identified the small molecule SynuClean-D as an attractive potential inhibitor of alpha-synuclein.
The compound could actively prevent alpha-synuclein aggregation in laboratory cells, with a reduction of Th-T levels between 34% and 58%, depending on the dose used. SynuClean-D could also reduce the amount of Th-T-positive aggregates in cells carrying the genetic mutations H50Q and A30P in the alpha-synuclein gene, which are associated with familial Parkinson’s disease.
Using a Caenorhabditis elegans worm model of Parkinson’s disease, the team found that treated animals had significantly fewer visible alpha-synuclein aggregates than untreated animals. In some worms, the treatment even resulted in a near-complete loss of protein aggregates.
“SynuClean-D is a nontoxic molecule that exhibits a unique capability to interact with and disassemble amyloid fibrils,” the researchers wrote.
Further analysis revealed that about 44% of treated animals retained all their dopamine-producing nerve cells alive, in contrast to only 14% of the untreated animals.
According to the researchers, these results “evidenced the ability of SynuClean-D to protect against alpha-synuclein–induced dopaminergic neuron degeneration,” a hallmark feature of Parkinson’s disease.
“Everything seems to indicate that the molecule we identified, the SynuClean-D, may provide therapeutic applications for the treatment of neurodegenerative diseases such as Parkinson’s in the future,” Salvador Ventura, PhD, a researcher at Universitat Autònoma de Barcelona and senior author of the study, said in a press release.
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Source: Parkinson's News Today