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Identifying serotonin receptors as a therapeutic target for SCA3

Written by Dr. Hannah K Shorrock Edited by Dr. Hayley McLoughlin

A C. elegans model of SCA3 helps to identify the serotonin receptor agonist befiradol as a potential therapeutic for SCA3

To move toward treatments for ataxia patients, it is important to understand what cellular and molecular pathways are dysfunctional in the disease. We can then identify specific cellular or molecular targets that are suitable for therapeutic intervention. In this study, researchers built on their previous work that adjusting serotonergic signaling could be used as a treatment approach in SCA3. They did this by identifying two drugs that target different combinations of serotonin receptors. When used together, these two drugs provide therapeutic benefit in Caenorhabditis elegans (C. elegans) models of SCA3. This study moves us closer to therapies for SCA3 by identifying the parts of the serotonergic signaling pathway most suitable to be targeted for further drug development.

The research team had previously worked with a small molecule that targeted multiple cellular pathways. This included many components of the serotonergic signaling pathway. In this study, the group wanted to identify the aspects of this signaling pathway responsible for the therapeutic benefit seen with these treatments. They used a C. elegans animal model of SCA3 in which a form of the SCA3 causing gene, ATXN3, containing 130 CAG repeats is expressed in neurons.

C. elegans have four receptors within the serotonergic signaling pathway that are highly similar to four receptors in humans. This makes them a strong model system to explore the treatment potential of targeting these receptors. The mutant ATXN3 C. elegans move less and they move slower than C. elegans without the ATXN3 expansion.

The group studied the effect of two drugs targeting receptors involved in the serotonergic signaling pathway on these movement defects. They found that at the same concentration both tandospirone and befiradol treatment for four days improved the motor performance of mutant ATXN3 C. elegans. This occurred for both the speed and amount of movement. Importantly, neither of the drugs affected the motor performance of C. elegans without the mutant ATXN3. This indicates that in this model system, the effect of the treatments is specific to the presence of the repeat expansion mutation. This is a desirable quality for any therapeutic strategy for a repeat expansion disease.

Two large blue worms on a black background, with three smaller blue worms beneath them
Microscopy images of C. elegans worms, a type of animal model used to study ataxia. Photo used under license by Heiti Paves/

What differentiated the efficacy of the two compounds was how quickly befiradol rescued motor performance. A short two-hour treatment with tandospirone did not improve motor performance for ATXN3 mutant C. elegans. However, a two-hour befiradol treatment completely restored the mutant C. elegans movement. This effect with befiradol was not just a short-term effect either. A two-hour treatment with befiradol improved motor performance for two days after the treatment had been removed. However, a longer treatment of befiradol (four days) did not extend the motor rescue after the treatment was removed. In both short (two-hour treatment) and long (four-day treatment), the therapeutic motor benefit was no longer seen by three days after befiradol was removed.  These results suggest two things. First, that befiradol and tandospirone may have different cellular targets. Second, the short versus long befiradol treatment may have slightly different effects on its targets.

Next, the group wanted to know if this improvement in motor performance was due to reducing the amount of aggregated mutant ATXN3 protein, as this is a hallmark of SCA3 disease. The group performed imaging studies and counted the number of ATXN3 protein aggregates. They also measured the total area of ATXN3 protein aggregates in mutant ATXN3 C. elegans. While neither short nor long treatment with tandospirone reduced the number or area of aggregates, befiradol was able to improve these phenotypes. The short, two-hour befiradol treatment reduced the number of aggregates. The longer four-day treatment reduced the total area of ATXN3 protein aggregates. This occurred without changing the overall levels of ATXN3 protein in the C. elegans. This is important as it indicates that befiradol treatment does not affect the healthy ATXN3 protein. Therefore should not hinder ATXN3 from performing its normal cellular functions.

The group wanted to identify the specific receptors within the serotonergic signaling pathway that are targeted by befiradol and tandospirone. While four serotonergic receptors are conserved between C. elegans and humans, there is at least one receptor in the serotonergic signaling pathway that is unique to the C. elegans genome. The group genetically removed each of these receptors individually in the ATXN3 mutant C. elegans. For two receptors, genetic removal did not influence the response of ATXN3 mutant C. elegans to befiradol or tandospirone. This indicates that these two receptors are not targeted by the treatments.

For another two of the receptors, there was no influence on the motor response to befiradol treatment. However, for these receptors, genetic ablation completely blocked any effect of tandospirone treatment on the motor phenotype in mutant ATXN3 C. elegans. These receptors are equivalent to human 5-HT6 and 5-HT7 receptors. The fifth receptor the group looked at had a large effect on the movement of mutant ATXN3 C. elegans. This receptor is equivalent to the human 5-HT1A receptor. Genetic ablation of this receptor improved the motor performance of mutant ATNX3 C. elegans while treatment with befiradol or tandospirone did not further improve motor performance. This shows that both treatments can act through this receptor. Together, this work demonstrates that befiradol acts on the 5-HT1A receptor equivalent while tandospirone can act on the 5-HT6, 5-HT7, and 5-HT1A equivalent receptors in SCA3 C. elegans models.

Finally, the group wanted to understand if befiradol directly interacts with 5-HT1A receptor equivalent in C. elegans. To do this, the group used a compound that directly blocks the binding site of the 5-HT1A receptor equivalent. By combining increasing concentrations of the blocking ligand with befiradol treatment, the group confirmed that it could prevent the befiradol-induced improvement in motor performance in a dose-dependent manner. This suggests that befiradol acts by directly binding to the ligand binding site of the 5-HT1A receptor equivalent in C. elegans. Together this work shows that treatments selective for binding to the 5-HT1A receptor ligand binding site may provide therapeutic benefit for SCA3 patients.

Tandospirone has previously been used in an ataxia clinical trial where it was shown to be safe and well-tolerated. During the 3 to 4 weeks of treatment, tandospirone improved the ataxia rating scales for SCA3 and SCA6 patients. This effect was however short-lived. Following the removal of tandospirone treatment, the improvement in symptoms was no longer present.

One possible reason for this is that tandospirone has multiple cellular targets, not just the 5-HT1A receptor, however, longer clinical trials have yet to be performed. It is therefore possible that a selective 5-HT1A receptor agonist may have greater therapeutic benefit for SCA3 patients. In clinical trials for other disorders, including painful peripheral diabetic polyneuropathy and movement disorders, befiradol was shown to be safe and well-tolerated but it has yet to be studied in ataxia patients. Future in-depth studies using mouse models will be the next step to validate befiradol use in SCA3, understand the role of the 5-HT1A receptor in the improvement of SCA3 symptoms, and ultimately move therapies targeting the serotonergic signaling pathway toward the clinic.

Key Terms

Receptor: Receptors are special structures that can receive and transduce a signal. In cell biology, they are made of protein and located within the cell membrane. When a receptor binds its signal molecule it is activated which leads to downstream effects. When the receptor is not bound to its signal molecule it is deactivated.

Caenorhabditis elegant (C. elegans): A type of very small worm-like animal called nematodes. C. elegans are very simple organisms, but can be used to learn about more diseases in more complex organisms like humans. To learn more, check out our Snapshot on C. elegans.

Conflict of Interest Statement

The author and editor declare no conflict of interest.

Citation of Article Reviewed

Pereira-Sousa, J., Ferreira-Lomba, B., Bellver-Sanchis, A., Vilasboas-Campos, D., Fernandes, J. H., Costa, M. D., Varney, M. A., Newman-Tancredi, A., Maciel, P. & Teixeira-Castro, A. Identification of the 5-HT1A serotonin receptor as a novel therapeutic target in a C. elegans model of Machado-Joseph disease. Neurobiol Dis 152, 105278, doi:10.1016/j.nbd.2021.105278 (2021).

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