Contribution and therapeutic potential of the cerebellum in amyotrophic lateral sclerosis – CEREBRALS

To overcome current limitations in understanding the motor circuits affected by ALS, the project leverages several innovative methods and technologies, combining studies in animal models and human patients.

1. Quantitative Computational Imaging

In humans, brain image analysis is performed using advanced magnetic resonance imaging (MRI) techniques. These methods enable precise visualization and measurement of cerebellar structures and their connections with other brain regions. The use of sophisticated computational tools allows for objective and reproducible quantification of the integrity of cerebellar lobules and cerebro-cerebellar connectivity pathways. This approach overcomes human interpretation bias and the limitations of classical analyses by offering high-resolution, unbiased mapping of anatomical and functional alterations.

2. Anterograde Viral Tracing in Mice

To explore the involved neural circuits in detail, viral tracing techniques are employed in mice. Specifically, modified, non-harmful viruses are injected into targeted cerebellar regions. These viruses act as “tracers”: they label neurons and allow researchers to track their projections and connections throughout the nervous system. Adeno-associated viruses (AAVs) are particularly well-suited for this purpose, as they selectively target neurons and can cross synapses, thereby revealing the connectivity pathways from the cerebellum to the spinal cord and other regions.

3. Gene Therapy Targeting the cerebellum

CEREBRALS explores the potential of gene therapy to correct neuronal dysfunctions observed in ALS. The gene therapy being developed aims to suppress the expression of the gene causing ALS within the cerebellum, with the goal of restoring normal function or compensating for the genetic abnormality. The effect of AAV injections, which silence the disease-causing gene in specific cerebellar regions, will be evaluated in terms of motor performance and lifespan in ALS mouse models.

4. Integration of Human and Animal Data

One of the major challenges in ALS research lies in bridging findings from animal models to humans. The project addresses this obstacle by applying similar analytical methods to both models: the results from viral tracing and functional analyses in mice are compared with human imaging data. This integrated approach enables validation of animal findings and identification of therapeutically relevant targets for humans.

Two original research articles analyze the contribution of the cerebellum to ALS and to disorders within the ALS–frontotemporal dementia (ALS-FTD) spectrum using advanced multimodal neuroimaging techniques.

In the study published in Neurology (2024), cerebellar involvement was longitudinally assessed in both sporadic and genetic forms of ALS. We demonstrate a progressive degeneration of cerebrocerebellar connections—particularly the cerebellofrontal and cerebelloparietal pathways—in sporadic patients, with early damage to the flocculonodular lobe and the cerebellar crura. Patients carrying C9orf72 mutations exhibited progressive atrophy of the ventral dentate nucleus. Alterations of the corticospinal and transcallosal tracts progressed faster than those of the cerebellar pathways, suggesting a progressive desynchronization between the cerebellum and the rest of the brain.

In the study published in the Journal of Neurology (2025), cerebello-cerebral connectivity profiles were investigated in patients with ALS-FTD, the behavioral variant of frontotemporal dementia (bvFTD), the non-fluent variant of primary progressive aphasia (nfvPPA), and the semantic variant of primary progressive aphasia (svPPA). A marked structural disconnection was observed between the cerebellum and the frontal, parietal, and temporal regions—particularly severe in nfvPPA. Spinocerebellar circuit damage and intra-cerebellar atrophy, especially affecting the posterior lobules and vermis, were also identified. Contrary to traditional views, cerebellar pathology significantly contributes to both motor and cognitive symptoms and cannot be overlooked in the ALS-FTD spectrum.

Together, these studies highlight that dysfunction of cerebellar networks -especially the deep nuclei and their projections- is a key and often underestimated component of ALS pathophysiology and its cognitive variants.

Our data, obtained through imaging, viral tracing, and functional tests in ALS mouse models, also show early defects in cerebellar pathways.

The CEREBRALS project opens up innovative research and development opportunities in the field of ALS by revealing the central role of the cerebellum in the disease’s pathophysiology. Through an integrative approach combining advanced human imaging, viral tracing, and behavioral studies in mice, several major avenues for application are emerging.

1. New Therapeutic Targets

The identification of descending circuits from the cerebellum to the spinal cord, and their early alteration in ALS, provides an as-yet unexplored anatomical target for gene therapy.

2. Innovative Imaging Biomarkers

Imaging analyses conducted in patients have highlighted a progressive degeneration of cerebello-cerebral circuits from the earliest stages. These alterations—correlated with motor and cognitive symptoms—could serve as diagnostic or prognostic biomarkers, with increased sensitivity compared to classic corticospinal markers.

3. Patient Care

The demonstration of the cerebellum’s contribution to motor (gait, coordination, dysarthria) and cognitive (behavior, language) disorders in ALS could influence patient management.

4. Expanded Preclinical Modeling

The mouse models used allow for the monitoring of early cerebellar alterations, including at presymptomatic stages. This creates a unique opportunity to reliably test the effectiveness of new therapies at an early stage.

5. Transferable Technological Tools

The protocol for trans-synaptic viral tracing combined with 3D imaging and cerebrospinal mapping is reproducible and could be applied to other neurodegenerative diseases involving motor coordination (spinocerebellar ataxias, Parkinson’s disease, etc.).

6. New Research Fields

Finally, the results renew interest in the concept of an ALS-ataxia continuum, particularly in relation to intermediate ATXN2 expansions. This could lead to a reclassification of clinical phenotypes, incorporating the cerebellar contribution into diagnostic criteria.

Amyotrophic lateral sclerosis (ALS) is an incurable neurodegenerative disease caused by the loss of motoneurons in the brain and spinal cord. Each year, ALS is responsible for three to five deaths per 100 000 people. ALS usually begins between the age of 50 and 70 with muscle weakness progressing rapidly to generalized paralysis and respiratory insufficiency that leads inexorably to death within 2 to 3 years of the onset of symptoms. Despite the wealth of knowledge on disease processes centered around the motoneuron, other lines of evidence suggest that ALS is multi-systemic and involves other regions of the central nervous system.
The consortium partners have recently uncovered structural defects in the cerebellum through clinical imaging studies in patients and observations in experimental models of the disease. This structure integrates sensory-motor information from the spinal cord, brainstem and cerebral cortex to regulate motor commands. Surprisingly, the involvement of the cerebellum has been largely underestimated in ALS even though its functions are central to the cardinal features of the disease. Our goal is to understand the role of the cerebellum in the disease and to evaluate its potential as a therapeutic target. This translational project involves neurobiologists and neurologists specialized in ALS and cerebellar development and function. The mission of CEREBRALS is to explore in patients and experimental models the dysfunctional neuroanatomical links between the cerebellum and other motor structures such as the spinal cord, using viral tracing and connectomics technologies by medical imaging and 3D tissue clearing. The therapeutic potential of the cerebellum will be validated by a viral transfer approach targeting the genetic cause of the pathology for proof of concept.