As for many other neurodegenerative diseases, there is no disease-modifying therapy for Spinocerebellar Ataxia Type 3 (SCA3) today [
1]. SCA3 is an autosomal-dominantly inherited neurodegenerative disease which is caused by an expansion of CAG-repeats in exon 10 of the
ataxin-3 (
Atxn3) gene leading consecutively to a higher number of glutamines (polyQ) in the ataxin-3 protein [
2,
3]. The polyQ repeats cause symptoms of ataxia if more than 50 glutamines are expressed [
4]. Even though this polyQ-expanded form of 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]. The physiological form of ataxin-3 protein is located in cell plasma whereas the polyQ-expanded protein is usually found both in cell plasma and nucleus and tends to form intranuclear aggregates in specific neurons during disease progression [
7]. Ataxia onset usually occurs at age 30–40 years [
1]. The clinical presentation, in addition to gait ataxia, dysarthria, and diplopia [
8,
9], is highly diverse and can present as parkinsonian phenotype [
10], sleep-associated movement disorder, restless legs syndrome [
11], or with psychiatric symptoms as depression [
12]. Genetic testing is the gold standard in diagnostics as the disease’s penetrance is 100% [
1,
13]. Up to now, the individual progression of disease is monitored clinically by ataxia scales like the Scale for the assessment and rating of ataxia (SARA) [
14]. However, 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 also a significant need for the investigation of easily accessible molecular biomarkers as surrogate endpoints for interventional trials testing new disease-modifying drugs [
15]. Therefore, it is crucial that newly established biomarkers are disease specific and that there is a correlation between surrogate and clinical endpoints [
16]. As for many other neurodegenerative diseases, neurofilament light chain (NfL) is shown to be increased in preataxic mutation carriers already 7.5 years before disease onset also in SCA3 even correlating with disease severity [
17‐
19]. However, NfL only reflects general axonal damage which captures disease progression and disease severity and, therefore, acts as a progression and severity biomarker. For ataxin-3 the main expected biomarker function would thus 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).
In this study, we, therefore, focused on the disease protein—ataxin-3—itself. As multiple targeted disease-protein lowering therapies in polyQ diseases like HD or SCA3 are currently being developed, there is a pressing need for sensitive (molecular) biomarkers for target engagement. For SCA3, it was previously shown that targeting the expanded
ataxin-3-allele by exon-skipping or
ataxin-3 gene suppression using antisense oligonucleotides (ASO) in SCA3 mouse models result in lower polyQ-expanded ataxin-3 levels and in a reduction of aggregation [
20‐
22]. Similar results are shown if the down-regulation of the disease protein ataxin-3 via specific micro-RNA (miRNA) is induced [
23‐
26]. To get prepared for ataxin-3 lowering therapies in further clinical trials, our study aimed to establish sensitive methods to measure total full-length and polyQ-expanded ataxin-3 protein in PBMCs, an easily accessible human biomaterial. Additionally, influencing factors as tube systems or the use of different sample processing protocols were validated to adapt highly standardized procedures.