Abstract
In view of upcoming clinical trials, quantitative molecular markers accessible in peripheral blood are of critical importance as prognostic or pharmacodynamic markers in genetic neurodegenerative diseases such as Spinocerebellar Ataxia Type 3 (SCA3), in particular for signaling target engagement. In this pilot study, we focused on the quantification of ataxin-3, the protein altered in SCA3, in human peripheral blood mononuclear cells (PBMCs) acquired from preataxic and ataxic SCA3 mutation carriers as well as healthy controls, as a molecular marker directly related to SCA3 pathophysiology. We established two different highly sensitive TR-FRET-based immunoassays to measure the protein levels of either total full-length, non-expanded and expanded, ataxin-3 or specifically polyQ-expanded ataxin-3. In PBMCs, a clear discrimination between SCA3 mutation carrier and controls were seen measuring polyQ-expanded ataxin-3 protein level. Additionally, polyQ-expanded ataxin-3 protein levels correlated with disease progression and clinical severity as assessed by the Scale for the Assessment and Rating of Ataxia. Total full-length ataxin-3 protein levels were directly influenced by the expression levels of the polyQ-expanded ataxin-3 protein, but were not correlated with clinical parameters. Assessment of ataxin-3 levels in fibroblasts or induced pluripotent stem cells allowed to distinguish mutation carriers from controls, thus providing proof-of-principle validation of our PBMC findings across cell lines. Total full-length or polyQ-expanded ataxin-3 protein was not detectable by TR-FRET assays in other biofluids like plasma or cerebrospinal fluid, indicating the need for ultra-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, the polyQ-expanded ataxin-3 protein is a promising candidate as a molecular target engagement marker in SCA3 in future clinical trials, determinable even in—easily accessible—peripheral blood biomaterials. These results, however, require validation in a larger cohort and further standardization of modifying conditions.
Highlights
As for many other neurodegenerative diseases, there is no disease-modifying therapy for Spinocerebellar Ataxia Type 3 (SCA3) today [1]
As SCA3 is a rare, autosomal-dominantly inherited neurodegenerative disorder with a slow disease progression, there is an urgent need for biomarker establishment to accomplish randomized clinical trials (RCT) [31], in particular of pharmacodynamics/ response biomarker which captures target engagement and is even accessible in peripheral blood
Our study demonstrates for the first time a systematic analysis of total full-length and polyQexpanded ataxin-3 protein levels across human biomaterials linked to clinical data
Summary
As for many other neurodegenerative diseases, there is no disease-modifying therapy for Spinocerebellar Ataxia Type 3 (SCA3) today [1]. The polyQ repeats cause symptoms of ataxia if more than 50 glutamines are expressed [4] Even though this polyQ-expanded form of Journal of Neurology (2021) 268:1304–1315 the ataxin-3 protein is expressed ubiquitously in somatic cells, selective neurodegeneration of deep cerebellar nuclei and basal ganglia is observed in SCA3 [5, 6]. Today there are no clear molecular biomarkers neither in cerebrospinal fluid (CSF), blood plasma, or serum nor in isolated blood cells like Peripheral Blood Mononuclear Cells (PBMC) to monitor disease onset or progression [15]. As the progression of SCA3 is slow, there is a significant need for the investigation of accessible molecular biomarkers as surrogate endpoints for interventional trials testing new disease-modifying drugs [15]. For ataxin-3 the main expected biomarker function would be classified as pharmacodynamics/ response biomarker which directly capture target engagement of the key disease protein (FDA classification based upon the context of the use of a biomarker, see https://www.fda.gov/drugs/biomarker-qualification-program/context-use)
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