2020
Gonsior K, Kaucher GA, Pelz P, et al. PolyQ-expanded ataxin-3 protein levels in peripheral blood mononuclear cells correlate with clinical parameters in SCA3: a pilot study. J Neurol 2021;268:1304–15
https://doi.org/10.1007/s00415-020-10274-y Abstract
In view of upcoming clinical trials in SCA3, quantitative molecular markers accessible in peripheral blood are of critical importance. In this pilot study, we focused on the quantification of ataxin-3, the disease protein of SCA3, in human peripheral blood mononuclear cells (PBMCs) acquired from pre-ataxic and ataxic SCA3 mutation carriers as well as healthy controls. Due to the CAG repeat mutation causing SCA3, ataxin-3 is expanded in SCA3. We established two different highly sensitive TR-FRET-based immunoassays to measure the protein levels of the normal and mutant, abnormally expanded ataxin-3. PBMC levels of expanded ataxin-3 protein clearly discriminated between SCA3 mutation carriers and controls. Additionally, levels of expanded ataxin-3 protein correlated with disease progression and clinical severity. In contrast, levels of normal ataxin-3 were not correlated with clinical parameters. Assessment of ataxin-3 levels in fibroblasts or induced pluripotent stem cells also allowed to distinguish mutation carriers from controls, showing that the findings in PBMCs findings are valid across cell lines. With our assay, ataxin-3 protein was detectable in cells, but not in biofluids like plasma or cerebrospinal fluid, indicating the need for more sensitive assays for these biofluids. Standardization studies revealed that tube systems, blood sampling, and PBMC preparation may influence ataxin-3 protein levels indicating a high demand for standardized protocols in biomarker studies. In conclusion, expanded ataxin-3 protein measured in PBMCs is a promising candidate as a molecular target engagement marker in SCA3 in future clinical trials. These results, however, require validation in a larger cohort and further standardization of modifying conditions.
Wilke C, Haas E, Reetz K, et al. Neurofilaments in spinocerebellar ataxia type 3: blood biomarkers at the preataxic and ataxic stage in humans and mice. EMBO Mol Med 2020;12:e11803
https://doi.org/10.15252/emmm.201911803 Abstract
With targeted treatments coming into reach for SCA3, easily accessible, validated biomarkers for clinical trials and drug development in animals are warranted. This is particularly important for the pre-ataxic disease stage, in which individuals who carry the mutation do not yet have symptoms. We assessed blood serum levels of neurofilament light (NfL) in ataxic and pre-ataxic subjects of ESMI and another multicentric SCA3 cohort, and in a SCA3 knock-in mouse model. NfL is a well-established marker of brain damage in various neurological diseases. Pre-ataxic and ataxic SCA3 subjects showed increased levels of NfL. In pre-ataxic subjects, NfL levels increased with proximity to the individual expected onset of ataxia, with significant NfL elevations already 7.5 years before onset. NfL levels correlated with disease severity and disease progression. Blood NfL increases in human SCA3 were paralleled by similar changes in SCA3 knock-in mice, here also starting before mice developed symptoms. In mice, NfL increase followed ataxin-3 aggregation and preceded loss of cerebellar Purkinje neurons. NfL might thus serve as an easily accessible and validated biomarker for SCA3. It is associated with earliest neuropathological changes, and may be used as a progression marker and a marker that indicates imminent onset of ataxia.
2021
Hengel H, Martus P, Faber J, et al. Characterization of Lifestyle in Spinocerebellar Ataxia Type 3 and Association with Disease Severity. Mov Disord 2022;37:405–10
https://doi.org/10.1002/mds.28844 Abstract
Lifestyle could influence the course of SCA3, but representative data are missing. The objective of this study was to characterize lifestyle in SCA3 and investigate possible associations with disease parameters. We collected data on smoking, alcohol consumption, physical activity, physiotherapy, and body mass index (BMI) from 243 patients with SCA3 and 119 controls from the ESMI cohort and tested for associations with age of onset, disease severity, and progression. Compared with controls, patients with SCA3 were less active and consumed less alcohol. Less physical activity and alcohol abstinence were associated with more severe disease, but not with progression rate or age at onset. Smoking, BMI, or physiotherapy did not correlate with disease parameters. Differences in lifestyle factors of patients with SCA3 and controls as well as associations of lifestyle factors with disease severity are likely driven by the influence of symptoms on behaviour. No association between lifestyle and disease progression was detected.
Hübener-Schmid J, Kuhlbrodt K, Peladan J, et al. Polyglutamine-Expanded Ataxin-3: A Target Engagement Marker for Spinocerebellar Ataxia Type 3 in Peripheral Blood. Mov Disord 2021;36:2675–81
https://doi.org/10.1002/mds.28749 Abstract
Although no curative therapy for SCA3 is yet available, preclinical gene silencing approaches that aim to reduce the levels of the mutant, abnormally expanded disease protein, ataxin-3, demonstrate promising results. In view of upcoming clinical trials, quantitative and easily accessible molecular markers are of critical importance as target engagement markers that indicate to which degree a given treatment reduces levels of mutant ataxin-3. We aimed at developing an ultrasensitive immunoassay to specifically measure mutant, abnormally expanded ataxin-3 in plasma and cerebrospinal fluid (CSF). Using the novel single molecule counting ataxin-3 immunoassay (Singulex), we analyzed cross-sectional and longitudinal biosamples from the ESMI cohort. Levels of mutant ataxin-3 were correlated with clinical parameters. Plasma concentrations of mutant ataxin-3 were stable over 1 year. In conclusion, this novel immunoassay is able to quantify mutant, abnormally expanded ataxin-3 in plasma and CSF. The assay is ready for use in clinical trials investigating gene silencing approaches for SCA3.
Faber J, Schaprian T, Berkan K, et al. Regional Brain and Spinal Cord Volume Loss in Spinocerebellar Ataxia Type 3. Mov Disord 2021;36:2273–81
https://doi.org/10.1002/mds.28610 Abstract
Given that new therapeutic options for SCA3 are on the horizon, there is a need for markers that reflect disease-related alterations, in particular, in the pre-ataxic stage. The objective of this study was to quantify regional brain and upper cervical spinal cord sizes in SCA3 with magnetic resonance imaging (MRI) across the entire time course of the disease. The study was performed in 210 ataxic and 48 pre-ataxic SCA3 mutation carriers and 63 healthy controls. Compared to healthy controls, the size of cervical spinal cord segments C3 and C2, medulla oblongata, pons, and pallidum, and the cerebellar anterior lobe was reduced in pre-ataxic SCA3 mutation carriers, that of cervical spinal cord segments C2 and C3, medulla oblongata, pons, midbrain, cerebellar lobules crus II and X, cerebellar white matter, and pallidum were reduced in ataxic SCA3 mutation carriers. Of all metrics studied, pontine volume showed the steepest decline across the disease course. It was correlated with ataxia severity, CAG repeat length, and age, and these factors explained almost 50% of the variance of pontine volume. In conclusion, regional brain and spinal cord tissue loss in SCA3 starts before ataxia onset. Pontine volume appears to be the most promising imaging biomarker candidate for interventional trials that aim at slowing the disease progression.
2022
Hengel H, Martus P, Faber J, et al. The frequency of non-motor symptoms in SCA3 and their association with disease severity and lifestyle factors. J Neurol 2023;270:944–52
https://doi.org/10.1007/s00415-022-11441-z Abstract
Non-motor symptoms (NMS) are a substantial burden for patients with SCA3. There are limited data on their frequency, and their relation with disease severity and activities of daily living is not clear. In addition, lifestyle may either influence or be affected by the occurrence of NMS. We therefore wished to characterize NMS in SCA3 and investigate possible associations with disease severity and lifestyle factors. We performed a cross-sectional analysis of NMS in 227 SCA3 patients, 42 pre-ataxic mutation carriers, and 112 controls of the ESMI cohort, and tested for associations with SARA score, activities of daily living, and the lifestyle factors alcohol consumption, smoking and physical activity. Sleep disturbance, restless legs syndrome, mild cognitive impairment, depression, bladder dysfunction and reduced vibration sense were frequent among SCA3 patients, while mainly absent in pre-ataxic mutation carriers. Except for restless legs syndrome, NMS correlated significantly with disease severity and activities of daily living. Patients with higher physical activity showed less cognitive impairment and fewer depressive symptoms, but these differences were not significant. This study revealed an association between disease severity and NMS, likely driven by the progression of the widespread disease process. Associations between lifestyle and NMS are likely due to the influence of NMS on lifestyle.
Maas RPPWM, Teerenstra S, Lima M, et al. Differential Temporal Dynamics of Axial and Appendicular Ataxia in SCA3. Mov Disord 2022;37:1850–60
https://doi.org/10.1002/mds.29135 Abstract
Disease severity in SCA3 is commonly defined by the Scale for the Assessment and Rating of Ataxia (SARA) sum score, but little is known about the contributions and progression patterns of individual items. In this study, we wished to investigate the temporal dynamics of SARA item scores in SCA3 and evaluate if clinical and demographic factors are differentially associated with evolution of axial and appendicular ataxia. SARA scores were longitudinally determined in 223 SCA3 patients of the ESMI cohort with a follow-up assessment after 1 year. An increase in SARA score was mainly driven by items related to gait, stance, sitting, and speech with a markedly smaller contribution of items related to arm and leg movements. Finger chase and nose-finger test scores showed the least deterioration at follow-up. Compared with the full set of SARA items, omission of both tests would result in lower sample size requirements for therapeutic trials. SARA progression was faster in men than in women. Despite considerable interindividual variability, the average annual progression rate of SARA score was approximately three times higher in subjects with a disease duration over 10 years than in those within 10 years from onset. Our findings will help inform the design of clinical trials and development of new outcome measures.
Ilg W, Müller B, Faber J, et al. Digital Gait Biomarkers Allow to Capture 1-Year Longitudinal Change in Spinocerebellar Ataxia Type 3 Mov Disord 2022;37:405–10
https://doi.org/10.1002/mds.29206 Abstract
With disease-modifying drugs on the horizon for SCA3, sensitive biomarkers that reflect motor function are highly warranted. Measures of step variability and body sway during gait have been shown to correlate with clinical ataxia severity in previous studies. However, to serve as valid progression biomarkers, these measures have to prove their capacity to capture longitudinal change, ideally within short time frames. We present the first multicentric longitudinal gait analysis study in SCA3. We performed a combined cross-sectional (n = 28) and longitudinal (1-year, n = 17) analysis in ataxic and pre-ataxic SCA3 mutation carriers from the ESMI cohort. Longitudinal analysis showed significant change in gait measures between baseline and 1-year follow-up, with high effect sizes. Sample size estimation for lateral sway indicated a required cohort size of n = 43 for detecting a 50% slowing down progression, compared with n = 240 with the Scale for the Assessment and Rating of Ataxia (SARA). These measures thus present promising motor biomarkers for upcoming interventional studies.
Garcia-Moreno H, Prudencio M, Thomas-Black G, et al. Tau and neurofilament light-chain as fluid biomarkers in spinocerebellar ataxia type 3. Eur J Neurol 2022;29:2439–52
https://doi.org/10.1111/ene.15373 Abstract
Clinical trials in SCA3 will require biomarkers for use as outcome measures. To evaluate total tau (t-tau), glial fibrillary acidic protein (GFAP), ubiquitin carboxy-terminal hydrolase L1 (UCHL1) and neurofilament light-chain (NfL) as fluid biomarkers in SCA3, we measured plasma concentrations of the four proteins on the Simoa HD-1 platform in 173 SCA3 mutation carriers and 172 controls of the ESMI cohort. Cerebrospinal fluid (CSF) samples of 11 mutation carrier were analysed for t-tau and phosphorylated tau (p-tau181). A transgenic SCA3 mouse model was used to measure cerebellar t-tau levels. Plasma t-tau levels were higher in mutation carriers below the age of 50 compared to controls. Pre-ataxic carriers showed higher CSF t-tau and p-tau181 concentrations compared to ataxic patients. Cerebellar t-tau was elevated in transgenic mice only in the early stages of the disease. GFAP and UCHL1 did not show higher levels in mutation carriers compared to controls. Plasma NfL concentrations were higher in mutation carriers compared to controls, and differences were greater for younger carriers. Our results suggest that tau might be a marker of early disease stages in SCA3. NfL can discriminate mutation carriers from controls and is associated with different clinical variables. Longitudinal studies are required to confirm their potential role as biomarkers in clinical trials.
2023
Santana MM, Gaspar LS, Pinto MM, et al. A standardised protocol for blood and cerebrospinal fluid collection and processing for biomarker research in ataxia. Neuropathol Appl Neurobiol 2023;49:e12892
https://doi.org/10.1111/nan.12892 Abstract
A major focus of ESMI is the identification of SCA3 biomarkers to enable future interventional studies. As the collection of biofluids and the way, how biofluids are processed, strongly impact the outcome of biomarker studies, we standardised biosampling procedures. Here, we describe the ESMI consensus biosampling protocol, developed within the scope of ESMI, that might be ultimately translated to other brain diseases, being the first step to protocol harmonisation in the field.
Raposo M, Hübener-Schmid J, Ferreira AF, et al. Blood transcriptome sequencing identifies biomarkers able to track disease stages in spinocerebellar ataxia type 3. Brain 2023;146:4132-43
https://doi.org/10.1093/brain/awad128 Abstract
Dysregulation of gene transcription is a known feature of SCA3. As ataxin-3 is expressed in the whole body, transcriptional alterations in blood may reflect early changes that start before clinical onset and might serve as biomarkers in clinical and research settings. Our goal was to describe dysregulated genes and cellular pathways, which can track disease onset, severity or progression in SCA3 mutation carriers. To identify dysregulated genes, were performed RNA sequencing of blood samples from 40 SCA3 mutation carriers and 20 healthy controls and compared the data with transcriptomic data from cerebella of autopsied SCA3 patients and controls. Ten genes - ABCA1, CEP72, PTGDS, SAFB2, SFSWAP, CCDC88C, SH2B1, LTBP4, MEG3 and TSPOAP1 - whose expression in blood was altered in the pre-ataxic stage and correlated with ataxia severity in the ataxia stage, were analysed by quantitative real-time PCR in blood samples from an independent set of 170 SCA3 mutation carriers and 57 controls. Pathway enrichment analysis indicated Gαi signalling and oestrogen receptor signalling were similarly affected in blood and cerebellum. SAFB2, SFSWAP and LTBP4 were dysregulated in pre-ataxic mutation carriers allowing to discriminate them from healthy controls with an accuracy of 79%. In SCA3 patients, ataxia severity was associated with higher levels of MEG3 and TSPOAP1. We propose expression levels of SAFB2, SFSWAP and LTBP4 as well as MEG3 and TSPOAP1 as stratification biomarkers, i.e. markers that help to assign SCA3 mutation carriers to more homogeneous subgroups.
Abstract
In view of the development of targeted therapies for SCA3, knowledge of early biomarker changes is needed. We analyzed cross-sectional data of 292 SCA3 mutation carriers from the ESMI cohort. Blood concentrations of mutant, abnormally expanded ataxin-3 protein were high before and after ataxia onset, whereas neurofilament light (NfL) was elevated and deviated from normal 13.3 years before onset. Pons and cerebellar white matter volumes measured with magnetic resonance imaging (MRI) decreased and deviated from normal 2.2 years and 0.6 years before ataxia onset. We propose a staging model of SCA3 that that includes an initial asymptomatic carrier stage followed by the biomarker stage characterized by absence of ataxia, but changes of NfL, as well as pons and cerebellar white matter volumes, finally leading into the ataxia stage, defined by manifest ataxia.
2024
Abstract
Cerebellar atrophy is the neuropathological hallmark of most ataxias. Hence, quantifying the volume of the cerebellar grey and white matter is of great interest. In this study, we aimed to identify volume differences in the cerebellum of SCA1, SCA3, and SCA6 as well as multiple system atrophy of cerebellar type (MSA-C). Cerebellar volumes were obtained from T1-weighted magnetic resonance images (MRI). Patients with MSA-C demonstrated the most pronounced volume loss, particularly in the cerebellar white matter. Patients with SCA6 showed more pronounced volume loss in cerebellar grey matter than to all other patient groups. MSA-C, SCA1 and SCA3 showed prominent atrophy of the cerebellar white matter. Our results characterize SCA6 as an almost pure cerebellar grey matter disease, emphasize the involvement of cerebellar white matter in SCA1, SCA3 and MSA-C, and reflect the rapid progression in MSA-C.
Raposo, M., J. Hubener-Schmid, R. Tagett, et al. Blood and Cerebellar Abundance of Atxn3 Splice Variants in Spinocerebellar Ataxia Type 3/Machado-Joseph Disease. Neurobiol Dis 2024;193:106456
https://doi.org/10.1016/j.nbd.2024.106456 Abstract
The gene causing SCA3, ATXN3, undergoes alternative splicing, i.e. a cellular process in which exons from the same gene are joined in different combinations leading to different isoforms of the encoded protein. Isoforms of the SCA3 disease protein, ataxin-3, differ in their toxicity, suggesting that certain ATXN3 splice variants may be crucial in driving the selective toxicity in SCA3. We sequenced RNA from blood (n = 60) and cerebellum (n = 12) of SCA3 mutation carriers and controls. The reference RNA transcript (ATXN3-251), translating into an ataxin-3 isoform harbouring three ubiquitin-interacting motifs (UIMs), showed the highest abundance in blood, while the most abundant transcript in the cerebellum (ATXN3-208) was of unclear function. Noteworthy, two of the four transcripts that encode full-length ataxin-3 isoforms but differ in the C-terminus had strong tissue specificity: ATXN3-251 (3 UIMs) was expressed in blood 50-fold more than in the cerebellum, whereas ATXN3-214 (2 UIMs) was expressed in the cerebellum 20-fold more than in the blood. These findings shed light on ATXN3 alternative splicing, aiding in the understanding of SCA3 pathogenesis and providing guidance in the design of future therapies.
Abstract
Little is known about the progression of health-related quality of life (HRQoL) and predicting factors in spinocerebellar ataxia (SCA). Such knowledge is crucial to identify modifiable factors promoting everyday life with SCA and attenuating HRQoL decline. To fill these knowledge gaps, longitudinal data (three-year follow-up) of 310 SCA patients of the ESMI and 525 SCA patients (SCA1, SCA2, SCA3 or SCA6) of the EUROSCA cohort (2006-2015) were assessed. HRQoL was assessed with the EQ-5D-3L, a questionnaire that asks for problems in mobility, self-care, usual activities, pain/discomfort and anxiety/depression. HRQoL significantly decreased over time. It was correlated with ataxia severity and depression. HRQoL more intensively declined in male and patients with an earlier age of onset. Faster progression of ataxia severity, depression, and overweight caused more severe decline of patients' HRQoL over time. In absence of curative treatments, stronger focus on mental health and body weight could help in improving HRQoL of SCA patients, especially in male patients with early disease onset.
Abstract
Health-related quality of life is a crucial outcome in clinical research. EQ-5D-3L is a widely used instrument to measure health-related quality of life. However, validation performance is lacking in patients with spinocerebellar ataxias. In this study, we analysed data from 842 SCA patients from the longitudinal ESMI and EUROSCA natural history studies. The objective was to assess the acceptability, validity, reliability, and responsiveness of the EQ-5D-3L in SCAs. The psychometric analysis of the data demonstrated that the EQ-5D-3L is an acceptable, valid, and reliable instrument for SCA patients and shows overall low responsiveness. Thus, the EQ-5D-3L could be used regularly in economic evaluations and as an additional instrument to the disease-specific HRQoL measures in SCA clinical research.
Abstract
In SCA3, first targeted gene therapy trials have started, offering the intriguing scenario of preventive treatment. SCA3 is associated with progressive regional brain atrophy that starts before clinical manifestation. We aimed to identify the progression pattern of brain atrophy of SCA3 with a focus on early disease stages. Magnetic resonance imaging (MRI) scans of 300 SCA3 mutation carriers and 317 controls were analyzed. A method called Subtype and Stage Inference (SuStaIn) was used to identify the sequence of volume loss across selected brain regions. We observed one distinct sequence of brain atrophy events in SCA3 without evidence for the existence of alternative cascades. Atrophy started in the most caudal parts of the brainstem. Almost all pre-ataxic SCA3 mutation carriers clustered in the first atrophy stages defined by SuStaIn. Certainty of sequence estimation was highest for early atrophy stages with prominent involvement of the pons and cerebellar white matter. Brain atrophy in SCA3 follows a clear and distinct sequence ascending from the lower brainstem with an early involvement of white matter. Knowledge of this sequence might support the identification of SCA3 mutation carriers with an imminent clinical onset for early interventions.
Elter T, Sturm D, Santana MM et al. Regional distribution of polymorphisms associated to the disease-causing gene of spinocerebellar ataxia type 3. J Neurol 2024;272:54
https://doi.org/10.1007/s00415-024-12829-9 Abstract
Genetic polymorphisms are small changes in the DNA sequence of a gene occurring between healthy individuals. Polymorphisms that are linked to the ATXN3 gene, especially to the pathological, expanded allele, are of particular interest for the development of a gene therapy for SCA3, because they may help to design compounds that specifically silence the expanded allele without interfering with the physiological, non-expanded allele. In this study, we investigated the frequency and regional distribution of two intragenic single nucleotide polymorphisms (SNPs) in the ESMI cohort. The distribution of genotypes on the expanded allele differed from those of the wildtype allele of SCA3 mutation carriers and of healthy controls, and was mainly influenced by the regional origin. In our cohort, no particular clinical phenotype was associated with these polymorphisms. Our results confirm distinct allocations of polymorphisms associated to the expanded ATXN3, and accordingly the consideration of allele-specific therapies.