‘Microsteretcher’ Technique May Advance Parkinson’s Research

microstretcher technique

Researchers have developed a way to closely study the function of a cell’s transportation network, made up of structures called microtubules, that appears to be impaired in people with Parkinson’s disease.

This technique, called “microstretcher,” may help to better understand microtubules’ deficits and their underlying causes in Parkinson’s, as well as to identify new therapeutic targets.

“Our experimental set-up enables us to study the relationship between the deformation of microtubules and their biological functions,” Akira Kakugo, PhD, the study’s senior author at Hokkaido University, in Japan, said in a press release.

The new method was described in the study, “Regulation of Biomolecular-Motor-Driven Cargo Transport by Microtubules under Mechanical Stress,” published in the journal ACS Applied Bio Materials.

Microtubules are hollow tubular structures that provide structure not only to cells, but also form highways to transport molecules and organelles inside cells. This transport is mediated by motor proteins, namely dynein and kinesin, which move in opposite directions along microtubules’ “tracks.”

In nerve cells, microtubules are particularly involved in the transport of vesicles filled with neurotransmitters (chemical messengers used in nerve cell communication) down the axon, or nerve cell fiber, to be released as signals to other nerve cells.

Increasing evidence suggests that defective regulation of microtubules may have a role in the development of a broad range of neurodevelopmental, psychiatric, and neurodegenerative diseases, including Parkinson’s.

Microtubule dysfunction was shown to precede axon transport deficits and death of dopamine-producing nerve cells — a hallmark of Parkinson’s disease. In addition, several Parkinson’s-associated proteins, such as tau, alpha-synuclein, parkinpink1, and LRRK2 regulate or appear to affect microtubule stability.

However, there was no experimental set-up to properly study the transport process in microtubules and the effects of disturbances in that process, until now.

Researchers at Hokkaido University and the National Institute of Information and Communications Technology, in Japan, developed a unique technique to control microtubular physical deformation and observe its effects on their transport function.

The microstretcher consists of a horizontal plate with flexible medium inside, which can be “stretched” or “compressed” using a computer system.

The team first used specific proteins to attach microtubules to the pre-stretched medium in a way that the microtubules lied parallel to the surface area and “stretching” axis. Next, dyneins bound to a fluorescent cargo were added to the microtubules.

With this system, researchers were able to evaluate the effects of stretching and compressing the flexible medium, and consequently straightening and bending the microtubules, in dynein-associated transport.

Results showed that the dynein-cargo moved faster as the microtubules began to bend, but only until the compressive strain reached about 25%, “beyond which the dynein-driven transport is retarded,” the researchers wrote.

From that point onward, the speed of dynein-associated transport started to decrease and eventually the deformation led to microtubule collapse and no dynein movement. Different speeds of motion also were observed along distinct areas of the bended microtubules.

The team noted that dynein’s faster motion in slightly bended, rather than straight, microtubules may be associated with the protein’s “walking-like” movement. Also, they believe these physical characteristics of microtubules may contribute to their functions in regulating many cellular processes.

“This work offers a technical advantage for a systematic study of the correlation between the deformation of MTs [microtubules] and its biological functions, i.e., cargo transport, as well as an opportunity to explore the interaction of deformed MTs with MT-associated proteins such as (…) tau,” the researchers wrote.

This new technique is “expected to help explain the [underlying disease-associated mechanisms] of traumatic brain injury, which mechanically stresses cells, and neurological conditions like Huntington’s and Parkinson’s diseases, in which microtubules are known to malfunction,” said Syeda Rubaiya Nasrin, the study’s first author.

Next, the team hopes to evaluate the effects of microtubule physical deformation in kinesin-driven transport along their “tracks.”

“The more we understand this process, the closer we might get to designing new nature-inspired materials that can act in a similar way,” Kakugo said.

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APOE Variant Directly Tied to Lewy Body Dementias in 2 Studies

APOE4 study

A variant of the apolipoprotein (APOE) protein, called APOE4, has been shown to directly affect Lewy body dementias, such as Parkinson’s disease.

Two separate studies, published simultaneously, found that APOE4 directly regulates levels of alpha-synuclein, which clumps  to form the nerve-damaging Lewy bodies that are the main culprits of the nerve cell death that defines Parkinson’s.

Their combined results help in understanding how APOE4 works, and how it affects disease progression. Greater insights into these mechanisms are vital for advancing research into treatments for Lewy body dementias.

Published in the peer-reviewed journal Science Translational Medicine, the two studies are “APOE4 exacerbates α-synuclein pathology and related toxicity independent of amyloid,” and “APOE genotype regulates pathology and disease progression in synucleinopathy.”

“It’s nice when you do science separately … but reach similar conclusions,” Guojun Bu, PhD, senior author of one study and chair of neuroscience at the Mayo Clinic, said in a news release published in Neurology Today.

APOE4 has been the focus of research into both Alzheimer’s and Parkinson’s for some time. Studies have shown that it strongly associates with these diseases, and that it plays a strong functional role in the accumulation of amyloid-beta and tau within neurons.

Whether APOE4 directly promotes alpha-synuclein aggregation or affects disease progression as a result of these aggregates, however, is not known.

In each of these studies, scientists engineered mice to express one of three APO variants — E2, E3, or E4 — or to have no APOE at all (knockout mice). They then used different methods to examine associations between the APOE variants and disease features, or pathology.

Albert Davis, an assistant professor of neurology at Washington University School of Medicine in St. Louis and colleagues monitored one group of each type of mice, looking for the development of alpha-synuclein aggregates. His group injected groups of each of these engineered mice with alpha-synuclein fibrils to induce protein clumping, and see how its spread varied in each genetic background.

Among the first group, those expressing APOE4 (E4) showed higher amounts of insoluble and phosphorylated (pathologic) alpha-synuclein, and evidence of reactive gliosis — a type of neuroinflammation — than did mice in other groups.

Reactive gliosis refers to inflammation of glial cells, a class of protective neurons that include microglia, a cell often seen to be damaged in Parkinson’s. This inflammation typically occurs in response to damage to the central nervous system (CNS), such as the formation of Lewy bodies.

Mice carrying the E2 variant survived longer and did not show the motor difficulties seen in the other mouse groups.

Among mice injected with alpha-synuclein fibrils to monitor its spread throughout the brain, the E4 mice showed the greatest signs of pathology within the substantia nigra, the brain region most affected by alpha-synuclein aggregates in Parkinson’s.

This finding closely matched that of another recent paper, which concluded that microglia play “an integral role in the propagation and spread of alpha-synuclein pathology.”

The two papers reached different conclusions, however, regarding the order of events in inflammation and alpha-synuclein/Lewy body formation. While Davis’s group concluded that alpha-synuclein pathology leads to an inflammatory response, the other research group, lead by Jeffrey Kordower of Rush University, concluded that inflammation came first and played a driving role in alpha-synuclein aggregation.

“We and others in the field are going to look closely at that and follow up,” Davis said in the release.

Davis’ group also examined the genetic background of two groups of Parkinson’s patients, as a comparison to the mouse models. His group found people that in both cohorts, those with two copies of the E4 variant, showed the fastest cognitive declines.

“Our results demonstrate that APOE genotype directly regulates alpha-synuclein pathology independent of its established effects on [beta amyloid] and tau, corroborate the finding that APOE e4 exacerbates pathology, and suggest that APOE e2 may protect against alpha-synuclein aggregation and neurodegeneration in synucleinopathies,” these researchers concluded in their paper.

In the second study, led by Bu at the Mayo Clinic, mice were injected with viruses carrying different APOE variants.

Similar to Davis’ study, Bu’s group found that mice expressing E4, but not E2 or E3, showed more alpha-synuclein pathology and Parkinson’s-related symptoms, such as impaired behavior and the loss of neurons and synapses (the junctions between neurons where information is passed from one nerve cell to another). The E4 mice also showed deficits in their fat and energy metabolism.

Gu and his colleagues examined the brains of patients with Lewy body dementia, and discovered that those who had the APOE4 variant also showed greater alpha-synuclein pathology.

Eric Reimann, the executive director of Banner Alzheimer Institute, praised the studies, while adding that their results need to be confirmed in larger groups of both Parkinson’s patients, “including those without comorbid (simultaneously occurring) Alzheimer’s disease,” and healthy controls.

When two or more medical co-existing conditions can be common, telling the effects of one apart from the other is challenging. This is especially the case in disorders such as Parkinson’s and Alzheimer’s, which share many of the same disease features.

Reiman had also found the E4 variant to associate with higher odds for Lewy body dementia. In contrast to Davis’ study, however, Reiman found no link between the E2 variant and a lower disease risk.

Alice Chen-Plotkin, an associate professor of neurology at the University of Pennsylvania Perelman School of Medicine, commented in the release that “the data for E4 being bad is much stronger than for E2 being good.”

Although she expressed surprise at the strength of the effect Davis’s group found APOE4 to have on glial cells, she noted that researchers are coming to think much more about these nervous system support cells.

An ongoing Phase 2 clinical trial (NCT04154072), for instance, seeks to improve Parkinson’s outcomes by blocking glial activation and inflammatory signaling. At the same time, the National Institutes of Health (NIH) recently awarded a $4.8 million grant to study how APOE4 induces neurodegeneration.

The E2 variant is also the focus of an ongoing Phase 1 gene therapy trial (NCT03634007), seeking to deliver this protein to patients’ CNS as a way of treating Alzheimer’s disease.

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Natural Killer Immune Cells Limit Parkinson’s Disease Progression, Study Finds

Natural killer cells

Immune cells known as “natural killer” (NK) cells appear to fight the cellular changes that lead to conditions such as Parkinson’s disease. Understanding how they do this may lead to new therapies for the neurodegenerative disorder, according to a study from the University of Georgia.

The study, “NK cells clear α-synuclein and the depletion of NK cells exacerbates synuclein pathology in a mouse model of α-synucleinopathy,” was published in the journal Proceedings of the National Academy of Sciences.

In Parkinson’s, patients progressively lose motor control and, to some degree, cognitive functions. One of the key clinical findings in Parkinson’s is the presence of insoluble clumps, or aggregates, of a protein called alpha-synuclein.

Alpha-synuclein is mainly found within the central nervous system (CNS), which is comprised of the brain and spinal cord. The clumps that build up in the brain are associated with the death of dopamine-producing (dopaminergic) neurons that lead to the disease’s symptoms.

They are one of the primary therapeutic targets among all Parkinson’s treatments. The discovery that the immune system can target these clumps on its own raises the possibility of designing better therapies based on the body’s own natural defenses.

Natural killer cells are a type of white blood cells that form the immune system’s first line of defense. They constantly patrol the body, attacking and destroying foreign bodies such as viruses and invading bacteria, as well as harmful native cells such as tumor cells.

Beginning with the observations that NK cells have been found to play a protective role in the CNS and that they are elevated in the blood of Parkinson’s patients, the UGA researchers sought to investigate their potential role in the disease.

Using mice, they looked closer at the relationship between NK cells and alpha-synuclein and experimented with the effect of removing NK cells in a mouse model of Parkinson’s.

As in human Parkinson’s patients, the researchers found natural killer cells in the brains of mice. There, these cells homed in on alpha-synuclein aggregates and showed evidence that they could absorb and degrade them. Importantly, the alpha-synuclein clumps reduced the NK cells’ cytotoxicity, with the killer cells releasing less of the pro-inflammatory — or inflammation-promoting — cytokine interferon-gamma, which is critical to both innate and adaptive immunity. Cytokines are small proteins released by immune cells that act upon other cells and have a specific effect on the interactions and communications between them.

Depleting NK cells in the mouse model caused Parkinson’s-like symptoms to progressively worsen, suggesting that natural killer cells did, in fact, exert a protective effect within the CNS of the mice.

Alongside the worsening of symptoms, the researchers observed that without the NK cells present, alpha-synuclein aggregates grew enlarged inside the mouse brains.

Interestingly, the depletion of natural killer cells in mice led to the death of dopaminergic neurons in the striatum of the brain, but not in the substantia nigra, where it usually occurs in humans. The striatum is a brain region responsible for the control of voluntary movement, while the susbtantia nigra contains a large percentage of all dopamine-producing neurons.

The study’s researchers point out that these results are preliminary and warrant more investigation into the role of NK cells in Parkinson’s disease.

“Our data suggest that NK cells have the potential to become the basis of a cell-therapeutic strategy to stop or slow abnormal protein pathogenesis in [Parkinson’s] and possibly other synuclein-related neurodegenerative diseases,” the researchers said.

Future steps will involve examining how natural killer cell function changes with age.

“Our preliminary data suggest that the number and function of NK cells are decreased in aged animals, and display impaired ability to perform their normal functions,” Jae-Kyung Lee, the study’s lead author, said in a press release.

“We would like to look deeper at age-related changes associated with NK cell biology and the wider implications for the health and well-being of older adults,” Lee said.

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Discovery of ‘Master Controller’ Region of Alpha-Synuclein Aggregation Opens Door to New Therapies

Alpha-Synuclein Aggregation

Scientists have discovered a “master controller” region that regulates aggregation of the alpha-synuclein protein, one of the distinctive features of Parkinson’s disease that is known to lead to the progressive degeneration of brain cells.

Understanding how the master controller works opens new opportunities in both the design of new therapies and in understanding the disease itself.

The findings were reported in the study, “A short motif in the N-terminal region of α-synuclein is critical for both aggregation and function,” published in the peer-reviewed journal Nature Structural and Molecular Biology.

Alpha-synuclein is found mainly in neurons, where, although its precise role remains unknown, it appears to be involved in neuron communication. However, in Parkinson’s, toxic clumps of alpha-synuclein — known as amyloid, or Lewy bodies — form, contributing to the onset and progression of the disease.

Previous studies have shown that a central region of the alpha-synuclein protein called NAC plays a key role in protein aggregation. However, less is known about the role other portions of the protein surrounding NAC may play in the process.

Now, researchers at the Astbury Centre for Structural Biology in the U.K. found two regions outside of NAC, called P1 and P2, that act as “master controllers” of alpha-synuclein aggregation.

In their experiments they showed that when they deleted or replaced both regions, alpha-synuclein was no longer able to form aggregates, even though NAC remained intact. Although these in vitro experiments established the biochemical importance of P1 and P2, further tests needed to be done to assess whether the same results could be found in living cells and organisms.

To that end, they inserted copies of alpha-synuclein lacking P1 and P2 into the muscle cells of the round worm C. elegans, a common animal model for neurological disorders.

After doing so, they found that while worms that had the normal, or wild-type version, of alpha-synuclein developed the typical toxic protein aggregates as they became older, animals lacking only the P1, or both master controller regions, did not.

These effects also were reflected in the worms’ behavior. Worms carrying the altered protein lacking either only P1 or both P1 and P2, were as mobile as healthy unmodified animals. In contrast, those carrying the version of alpha-synuclein with both master controller regions intact, showed age-dependent declines in motility.

“Our discovery of master controller regions may open up new opportunities to understand how mutating the protein sequence that causes disease could help us find the Achilles heel for these proteins to target for future therapeutic intervention,” Sheena Radford, PhD, co-author of the study said in a press release. Radford is director of the Astbury Centre for Structural Molecular Biology at the University of Leeds.

During neurotransmission, chemical messengers called neurotransmitters are wrapped up in a balloon-like membrane (vesicle), which then fuses with the nerve cell membrane in order to release its contents and propagate the message. Alpha-synuclein is thought to facilitate this process of membrane fusion. (Neurotransmission is the process by which neurons communicate with each other.)

Since P1 and P2 are both located in a region of alpha-synuclein that is responsible for interacting with cell membranes, researchers then wondered what the impact would be of eliminating these regions on alpha-synuclein’s ability to facilitate signaling between neurons.

They found that removing these master control regions interfered with the normal process of membrane fusion, impairing neurotransmission.

So, attempts to use the master control region therapeutically will have to balance the beneficial effects of preventing alpha-synuclein aggregation and the negative effects of compromising neuron communication.

“Our hope is that future research might target this master controller, to allow the development of a therapy which could tweak the conformation or stickiness of alpha-synuclein in the brain with only minimal changes to its function,” said David Brockwell, PhD, also a co-author of the study.

“We hope that such a strategy might be able to help people with early signs of Parkinson’s, by reducing the formation of amyloid plaques in the brain, and to delay the progression of the disease,” Brockwell said.

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Sangamo, Biogen Team Up to Develop Gene Therapies for Neuromuscular Diseases

gene therapies collaboration

Sangamo and Biogen are teaming up to develop gene therapies for Parkinson’s and Alzheimer’s disease, as well as another neuromuscular disease target and up to nine undisclosed neurological disorders.

The collaboration will leverage Sangamo’s proprietary zinc finger protein (ZFP) technology, designed to target almost any sequence in the human genome, which is the complete set of DNA. Delivered via a modified, harmless form of a virus, called an adeno-associated virus (AAV), the therapy modulates — either repressing or activating — the expression of key genes involved in neurological diseases.

Gene expression is the process by which information in a gene is synthesized to create a working product, like a protein.

“As a pioneer in neuroscience, Biogen will collaborate with Sangamo on a new gene regulation therapy approach, working at the DNA level, with the potential to treat challenging neurological diseases of global significance. We aim to develop and advance these programs forward to investigational new drug applications,” Alfred Sandrock Jr., MD, PhD, executive vice president, research and development at Biogen, said in a press release.

“There are currently no approved disease modifying treatments for patients with many devastating neurodegenerative diseases such as Alzheimer’s and Parkinson’s, creating an urgency for the development of medicines that will not just address symptoms like the current standards of care, but slow or stop the progression of disease,” said Sandy Macrae, CEO of Sangamo.

“We believe that the promise of genomic medicine in neuroscience is to provide a one-time treatment for patients to alter their disease natural history by addressing the underlying cause at the genomic level,” Macrae said.

ZFPs are part of Sangamo’s genome regulation technology. Delivered using AAVs, ZFPs can selectively activate or repress specific genes. In the case of Parkinson’s, ST-502 delivers ZFPs that target and repress the activity of the alpha-synuclein (SNCA) gene, which codes for the alpha-synuclein protein. Abnormal aggregates, or clumps of this protein are thought to underlie the development of Parkinson’s disease.

ST-501 works under the same principle but targets the protein tau, thought to be a key player in the development of Alzheimer’s disease.

In preclinical studies, strategies using these gene therapies have shown promising therapeutic effects.

“The combination of Sangamo’s proprietary zinc finger technology, Biogen’s unmatched neuroscience research, drug development, and commercialization experience and capabilities, and our shared commitment to bring innovative medicines to patients with neurological diseases establishes the foundation for a robust and compelling collaboration,” said Stephane Boissel, head of corporate strategy at Sangamo.

“This collaboration exemplifies Sangamo’s commitment to our ongoing strategy to partner programs that address substantial and diverse patient populations in disease areas requiring complex clinical trial designs and commercial pathways, therefore bringing treatments to patients faster and more efficiently, while deriving maximum value from our platform,” Boissel added.

Under the terms of the collaboration, Biogen has the exclusive global rights for ST-502, ST-501, and a third undisclosed neuromuscular disease target.

Moreover, Biogen has the exclusive rights to select up to nine additional undisclosed targets over a period of five years.

Sangamo will be responsible for early research for these gene therapies, with the costs shared by the companies. Biogen will take over research aimed at clinical development, regulatory interactions, and global commercialization, assuming responsibility for such costs.

Within the licensing agreement, Biogen will pay Sangamo $350 million upfront. Sangamo is eligible to receive up to $2.37 billion in potential milestones, as well as royalties once the therapies are marketed.

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New Cellular Model Can Reconstruct Entire Process of Lewy Body Formation in Parkinson’s Disease, Study Says

Lewy body formation, Parkinson's

Scientists have developed a new cellular model that can help reconstruct the entire process of Lewy body formation — a key event that underlies neurodegeneration in Parkinson’s disease — and that could potentially be used to evaluate the effect of therapeutics on the toxic protein buildup observed during this process.

The study, “The process of Lewy body formation, rather than simply α-synuclein fibrillization, is one of the major drivers of neurodegeneration,” was published in PNAS.

Parkinson’s is a multisystem neurodegenerative disorder with motor and non-motor features caused by the death of midbrain dopamine-producing neurons. These nerve cells are thought to die as a consequence of the aggregation, or clumping together, of a protein called alpha-synuclein in small fiber-like, or insoluble fibril, structures known as Lewy bodies.

Although evidence indicates abnormal alpha-synuclein accumulation is essential for the development of Parkinson’s disease, not much is known about the molecular and cellular processes that control the transformation of healthy alpha-synuclein protein into insoluble fibrils and, consequently, their clumping into Lewy bodies.

To better understand these biological events at a genetic, molecular, biochemical, structural, and cellular level, researchers at the Brain Mind Institute in Lausanne, Switzerland, tracked the development of Lewy bodies from beginning to end.

They began by using mouse primary neurons grown in lab dishes. They then added a small amount of alpha-synuclein fibrils that would be used as “seeds” that grew by recruiting neuron-produced alpha-synuclein.

“This approach has proven incredibly useful in modeling the formation of alpha-synuclein aggregates linked to diseases like Parkinson’s,” study senior author Hilal Lashuel, PhD, said in a press release.

Scientists usually monitor cell cultures for two weeks, but Lashuel’s team decided to go beyond that time limit. Twenty-one days after seeding, the team observed Lewy body-like inclusions in approximately 22% of the neurons.

These lab-grown Parkinson’s-related structures shared between 15%-20% of their protein content with “natural” Lewy bodies, which, given the slower growth rate in the laboratory versus the human brain, is an encouraging finding.

Importantly, the Lewy-body-like clump structures had not appeared by the usual two-week cutoff point, highlighting the need to prolong the experiments to observe features characteristic of neurodegeneration.

“Time, being patient and using a swiss army knife analytical approach is all that was required,” Lashuel said.

The formation of Lewy bodies involved a complex interplay between alpha-synuclein fibrillization, protein modifications, and interactions between alpha-synuclein clumps and membranous organelles, including mitochondria, which produce energy for cells.

In addition, the researchers found that Lewy body formation — rather than simply alpha-synuclein fibril accumulation — were the major drivers of neurodegeneration, since these structures disrupted cellular functions such as energy production, and compromised nerve cell communication by changing the properties of synapses, or the junctions between nerve cells that allows them to communicate.

Formation of Lewy bodies does not occur simply through the continued formation, growth, and assembly of alpha-synuclein fibrils “but instead arises as a result of complex [alpha-synuclein] aggregation-dependent events that involve the active recruitment and sequestration of proteins and organelles over time,” the researchers wrote.

“Our results generally agree with recent findings reported on Lewy bodies from Parkinson’s disease brains,” Lashuel said. “But while previous studies only offered snapshots of fibril evolution, our model can reconstruct the entire process of Lewy body formation, making it a powerful platform for elucidating the relationship between fibrillization and neurodegeneration in Parkinson’s and other diseases, and to screen for novel potentially disease modifying therapies.”

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Study Shows Age-dependent Spread Of Alpha-synuclein From Gut to Brain in Mice

gut Parkinson's alpha-synuclein

Aggregates of the Parkinson’s disease-associated protein alpha-synuclein can spread from the gut to the brain in mice, but this process is dependent on aging, likely as a result of altered protein dynamics, a new study suggests.

The study, “Gut-seeded α-synuclein fibrils promote gut dysfunction and brain pathology specifically in aged mice,” was published in Nature Neuroscience.

Parkinson’s disease is characterized by aggregates of alpha-synuclein in the nervous system, particularly in dopamine-producing neurons in the brain. However, how these aggregates form in the first place isn’t fully understood.

One proposed hypothesis is that these aggregates don’t initially form in the brain, but in the enteric nervous system (ENS) — the nervous system of the gut. This hypothesis is supported by the fact that many people with Parkinson’s experience gastrointestinal symptoms, such as constipation, years before the onset of motor symptoms. The idea is that, from the gut, the aggregates could “spread up” to the brain through the vagus nerve.

“The vagus nerve is a physical connection between neurons in the gut and neurons in the brain,” study co-author Collin Challis, PhD, a former postdoctoral researcher at California Institute of Technology (Caltech), said in a press release. “If these damaging protein clusters first originate in gut neurons, then in the future we may be able to diagnose [Parkinson’s disease] earlier and potentially use gene delivery to restore functions to the cells so that they can clean up the aggregates.”

In the new study, researchers injected alpha-synuclein aggregates into the lining of the intestinal tract in mice. This led to numerous Parkinson’s-associated changes in the gut, including increased production of inflammatory molecules (e.g. IL-6), gastrointestinal dysfunction, and increased activation of neurons in the ENS, indicating an inflammatory response.

The researchers then looked to see whether the aggregates had spread into the central nervous system (CNS, comprising the brain and spinal cord). Interestingly, initial experiments did not reveal significant aggregate accumulation in the CNS, nor were there sustained motor impairments of the sort that would be expected if this system was modeling Parkinson’s disease.

But, crucially, these experiments were done in young adult mice (8-10 weeks old), and the biggest risk factor for developing Parkinson’s is aging. Thus, the researchers repeated the experiment in older (16-month-old) mice.

In these mice, as in the younger mice, injecting alpha-synuclein into the gut lining led to gut dysfunction. But, unlike in the young mice, older mice also developed motor deficits resembling Parkinson’s disease. Furthermore, the older mice did have alpha-synuclein aggregates in their brainstems, supporting the idea that the aggregates “spread up.”

Additionally, the older mice had significantly reduced levels of dopamine in their brains, which was not observed in the younger mice following alpha-synuclein injection.

The reason for this age-related difference may come down to how protein-regulating systems in the body change with age.

The researchers demonstrated that older mice express significantly less of the gene GBA1, which encodes for the protein glucocerebrosidase (GCase). This protein plays a critical role in the molecular machinery that cells use to recycle proteins, so the researchers proposed that the age-associated decrease in GCase levels could allow alpha-synuclein aggregates to spread in a way that is limited in younger animals.

In keeping with this idea, the researchers found that, when they increased GCase levels in the guts of young adult mice through gene therapy using a variant of adeno-associated virus (AAV) as a delivery system, gut-related problems associated with alpha-synuclein were diminished, though not entirely resolved. As such, although GCase dysfunction probably isn’t the only factor, it likely plays a role in alpha-synuclein pathology.

“Our results propose mechanisms that may underlie the etiology of sporadic [Parkison’s disease] and highlight GBA1 as a therapeutic target for prodromal [early], peripheral synucleinopathy,” the researchers wrote.

“Interestingly,” they noted, “[disease-causing alpha-synuclein] also inhibits GCase function.” As such, it’s possible that there is a feedback loop wherein increasing alpha-synuclein leads to decreased GCase function, which in turn leads to further increases in alpha-synuclein, until a threshold is crossed, spilling over into disease.

“Mutations in the gene that encodes GCase are responsible for Gaucher disease and a risk factor in [Parkinson’s disease],” said Viviana Gradinaru, PhD, a professor at Caltech and co-author of the study. “Our work shows that this gene can be delivered by AAVs to rescue gastric symptoms in mice, and emphasizes that peripheral neurons are a worthwhile target for treating [Parkinson’s disease], in addition to the brain.”

Overall, the researchers said, “[O]ur findings suggest that age-related declines in protein homeostasis, including diminished GCase function, may promote susceptibility to [disease causing alpha-synuclein] in the ENS and support the gut-to-brain hypothesis of [the biology of] synucleinopathy.”

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APOE Gene Variants Alter Alpha-synuclein Dynamics, Could Affect Dementia Occurrence in Parkinson’s, Study Suggests

APOE Gene Variants

Genetic variations in the gene apolipoprotein E (APOE) alter the dynamics of alpha-synuclein protein buildup in the brains of mice with Parkinson’s disease (PD), according to a new study. This suggests suggests that alterations in APOE could affect the occurrence of dementia in humans with the neurodegenerative disease, the researchers said.

Titled “APOE genotype regulates pathology and disease progression in synucleinopathy,” the study was published in Science Translational Medicine.

As many as 80% of people with PD will develop dementia — a group of symptoms affecting memory, thinking and social abilities — within two decades of being diagnosed. Nonetheless, the occurrence of dementia in Parkinson’s varies greatly person-to-person: “Many patients take years to develop dementia, whereas others have a more rapid course, and in some cases, dementia precedes motor symptoms,” the researchers said.

Parkinson’s is associated with the formation of toxic protein aggregates, or clumps, in the brain — particularly involving the protein alpha-synuclein. Alzheimer’s disease also is characterized by the accumulation of toxic protein aggregates in the brain, though the exact proteins involved are somewhat different.

The gene APOE encodes a protein of the same name, which helps form molecules called lipoproteins, which are responsible for packaging cholesterol and other fats and carrying them through the bloodstream.

A variant in this gene, called E4, is associated with a significantly increased risk of Alzheimer’s. APOE variants are known to affect how certain Alzheimer’s-associated proteins clump together in the brain. However, whether these variants also affect alpha-synuclein aggregation hasn’t been clear, nor has been the effect of variants in APOE on dementia in Parkinson’s.

Now, researchers at Washington University School of Medicine in St. Louis are trying to bridge this knowledge gap. Using mice with a form of alpha-synuclein prone to aggregation, the scientists engineered mice with one of three genetic variants in APOE — E2, E3, or E4 — or no APOE gene at all, called a knockout. They then compared alpha-synuclein in the brains of these mice.

Using multiple molecular assays, the researchers demonstrated that alpha-synuclein levels were significantly lower in mice with the E2 variant than in mice with the E4 variant or knockout mice. Animals with the E3 variant had alpha-synuclein levels in between these two extremes, though differences were generally not statistically significant in either direction.

Motor function and survival patterns followed trends consistent with this finding: E2 mice had higher motor scores, followed by E3, then E4, and knockout. Similarly, E2 mice survived significantly longer (median 18.4 months) than E4 (11.7 months) or knockout mice (11.6 months), with E3 mice in between the extremes (median 12.7 months).

These data suggest that APOE genetic variants affect the dynamics of alpha-synuclein in the brain.

“What really stood out is how much less affected the APOE2 mice were than the others,” Albert (Gus) Davis, MD, PhD, a professor at Washington University School of Medicine and the study’s lead author, said in a press release.

“It actually may have a protective effect, and we are investigating this now,” Davis said. “If we do find that APOE2 is protective, we might be able to use that information to design therapies to reduce the risk of dementia.”

The team then looked for connections between APOE variants and dementia in people with Parkinson’s.

First, the researchers assessed two groups of PD patients who had been followed for several years: one from the Parkinson’s Progression Markers Initiative, involving 251 people, and the other, which includes 170 people, from the Washington University Movement Disorders Center. In both groups, individuals with the E4 variant had significantly faster rates of decline in cognitive scores as compared with those with other variants. Importantly, this effect remained significant after adjustment for other factors known to affect cognitive decline, including the presence of neurotoxic proteins in the fluid around the brain, and educational attainment.

Additionally, in a separate group — the NeuroGenetics Research Consortium, numbering 1,030 people, in which cognitive scores were measured at only one time point — the E4 variant was associated with significantly lower cognitive scores at the time of assessment, and with the onset of cognitive difficulties at a younger age.

“Together, these data corroborate the finding that APOE ε4 is associated with cognitive impairment and a faster rate of cognitive decline in PD,” the researchers said.

Because this effect was independent of other toxic brain proteins, the team concluded this most likely was a consequence of increased alpha-synuclein in the brain, as evidenced by the data in mice.

All in all, these findings implicate APOE in the molecular progression of Parkinson’s and, specifically, the onset of dementia. Thus, APOE or related proteins in the brain might be a viable therapeutic target for treating dementia in Parkinson’s, the researchers said. Further studies will be needed to test this idea.

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Structural Differences Seen in Toxic Proteins Marking Parkinson’s, Multiple System Atrophy

alpha-synuclein structure

Assessing the shape of alpha-synuclein aggregates in the brain helps to distinguish between Parkinson’s disease (PD) and another progressive neurodegenerative disorder known as multiple system atrophy (MSA), a study suggests.

The study, “Discriminating α-synuclein strains in Parkinson’s disease and multiple system atrophy,” was published in Nature.

Parkinson’s and MSA share many symptoms, particularly in earlier disease stages. This often leads to misdiagnoses, which pose an obstacle for early treatment.

Both PD and MSA are also characterized by the buildup of the protein alpha-synuclein in the brain, which is toxic to brain cells.

Researchers analyzed alpha-synuclein aggregates in cerebrospinal fluid (that which surrounds the brain and spinal cord) taken from 94 people with Parkinson’s, 75 with MSA, and 56 individuals with other neurological diseases serving as controls.

They used a technique called Protein Misfolding Cyclic Amplification (PMCA). This assay involves cyclic chemical reactions that make more of the protein — similar to a ‘copying’ technique called polymerase chain reaction (PCR) used with DNA.

Fluorescent dyes can then be used to gain insights into the structure of the proteins. Basically, the dyes bind to the proteins differently depending on their shape, which affects how they glow.

Several notable differences were evident between the fluorescence profiles of alpha-synuclein aggregates taken from people with MSA and those with PD. In MSA samples, fluorescence tended to increase quickly to a plateau of less than 1,800 fluorescent units. Parkinson’s samples, in contrast, tended to increase more slowly but reach a higher plateau — between 2,000 and 8,000 fluorescent units. Control samples did not show any fluorescence over the background levels.

Based on the results of this test, 85/88 (96.6%) PD samples and 61/65 (93.8%) MSA samples could be correctly identified as such. Some samples did not form aggregates at all, resulting in no fluorescent reading.

“Combining all samples, we correctly distinguished PD from MSA in 146 of the 153 samples analyzed—an overall sensitivity of 95.4%,” the researchers wrote.

Importantly, these same differences in fluorescent profiles were also observed in alpha-synuclein in brain tissue samples from people who had died with either disease. This suggests structural differences are inherent in the alpha-synuclein aggregates that form in the brains of people with Parkinson’s and with MSA.

Further analysis verified this finding. The researchers used a number of molecular tests to tease out the differences in shapes between the two types of alpha-synuclein. They also performed assays to confirm that this result was due to the shape of the protein, not the amount of protein in different samples or other factors.

Their work demonstrated that different fluorescent dyes bound to PD or MSA alpha-synuclein in different ways, indicative of their structural differences. The researchers also used cryo-electron microscopy to more directly assess the ‘shape’ of the proteins.

“[Alpha-synuclein] filaments from patients with MSA are predominantly twisted, whereas those from patients with PD are mostly straight,” the researchers wrote.

“It is important for physicians to have an objective way to differentiate between PD and MSA in order to provide patients with the best care. Currently, the only way to differentiate them is to wait and see how the disease progresses, with MSA advancing much more rapidly than PD,” Claudio Soto, PhD, the study’s senior author and a neurology professor with The University of Texas Health Science Center at Houston (UTHealth), said in a press release.

“By the time people show progressed symptoms of MSA, a substantial amount of brain cells are already damaged or dead, and they can’t be brought back. It has been difficult to develop treatment for both diseases because of the high rates of misdiagnosis, so we needed to find a way to distinguish between the two at the onset of early symptoms,” Soto added.

These structural differences likely have important biological consequences, which the researchers assessed by exposing cells in dishes to alpha-synuclein aggregates derived from people with MSA or PD. The MSA aggregates were significantly more toxic to the cells, possibly as a consequence of their different structures.

“Our results demonstrate that α-syn [alpha synuclein] aggregates exist as distinct conformational strains with different biochemical and structural properties, which will help to improve our understanding of the pathogenesis of these diseases,” the researchers concluded.

“These data may enable the development of a biochemical test for the specific diagnosis of different disorders that involve the misfolding of α-syn, with potential future applications in clinical trials and personalized medicine,” they added.

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Seelos Set to Begin Animal Studies for SLS-007 Using Gene Therapy Approach


Seelos Therapeutics will begin animal studies to test a gene therapy approach to deliver its investigational candidate SLS-007 for Parkinson’s disease.

SLS-007, originally developed by scientists at the University of California, Los Angeles, is intended to lessen the aggregation, or clumping, of alpha-synuclein protein — a major component of the Lewy bodies found in Parkinson’s patients.

The investigational therapy consists of a family of peptide blockers that specifically target a central region of alpha-synuclein called the non-amyloid component core, which is particularly prone to aggregation.

In preclinical studies, SLS-007 was able to slow disease progression, including stopping alpha-synuclein propagation and seeding — a process in which a “seed” provides the template for the aggregation of normal protein into clumps. This occurred when scientists used fibril preparations and alpha-synuclein seeds collected from patients with Parkinson’s or Lewy body dementia.

Now, the researchers at Seelos will test a modified, harmless form of a virus, called an adeno-associated virus, or AAV, as a vehicle to deliver SLS-007. In preparation, the company currently is producing the viral vectors and preparing the animals for its preclinical study.

“Identifying the AAV vector as the optimal route of systemic administration of SLS-007 may ultimately yield a convenient one-time delivery of the peptides,” Raj Mehra, PhD, chairman and CEO of Seelos, said in a press release.

“This is a novel method of viral delivery that has been a collaboration between our R&D team and key opinion leaders that, if successful, could be a major advancement in the field,” Mehra said.

The aim of the animal studies is to test the best route of administration, establish the pharmacokinetic and pharmacodynamic profiles of SLS-007, and assess parameters of target engagement to alpha-synuclein. In pharmacokinetics, researchers evaluate the movement of a compound into, through, and out of the body —essentially how the body affects a medicine. Pharmacodynamics determines the interactions between the body and a compound.

“The study will include measurements of several key biomarkers that will assess the extent of alpha-synuclein (α-synuclein) aggregates expression in key target areas of the brain, such as the forebrain and the substantia nigra,” said Tim Whitaker, MD, Seelos’ head of research and development.

“These outcomes will help determine key target engagement and set the stage for our next steps with the program,” he added.

The results from these studies, namely delivery and target engagement, are expected in the second half of this year.

Seelos also is developing SLS-004, its other candidate to treat Parkinson’s disease. SLS-004 uses another modified, harmless form of a virus, known as a lentivirus, to deliver an enzyme called DNA methyltransferase 3A. This targets a particular region of the SCNA gene, which provides the instructions for making alpha-synuclein, and limits the protein’s production.

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