Antisense Oligonucleotide-Mediated Removal of the Polyglutamine Repeat in Spinocerebellar Ataxia Type 3 Mice

Abstract

Spinocerebellar ataxia type 3 (SCA3) is a currently incurable neurodegenerative disorder caused by a CAG triplet expansion in exon 10 of the ATXN3 gene. The resultant expanded polyglutamine stretch in the mutant ataxin-3 protein causes a gain of toxic function, which eventually leads to neurodegeneration. One important function of ataxin-3 is its involvement in the proteasomal protein degradation pathway, and long-term downregulation of the protein may therefore not be desirable. In the current study, we made use of antisense oligonucleotides to mask predicted exonic splicing signals, resulting in exon 10 skipping from ATXN3 pre-mRNA. This led to formation of a truncated ataxin-3 protein lacking the toxic polyglutamine expansion, but retaining its ubiquitin binding and cleavage function. Repeated intracerebroventricular injections of the antisense oligonucleotides in a SCA3 mouse model led to exon skipping and formation of the modified ataxin-3 protein throughout the mouse brain. Exon skipping was long lasting, with the modified protein being detectable for at least 2.5 months after antisense oligonucleotide injection. A reduction in insoluble ataxin-3 and nuclear accumulation was observed following antisense oligonucleotide treatment, indicating a beneficial effect on pathogenicity. Together, these data suggest that exon 10 skipping is a promising therapeutic approach for SCA3.

Oligonucleotide therapy mitigates disease in spinocerebellar ataxia type 3 mice

“…These results further support the growing evidence from preclinical animal and human clinical ASO trials demonstrating sustained, widespread, and well-tolerated ASO delivery to the CNS following longitudinal ICV or intrathecal delivery 20,22 . In support of the tolerability of ASOs for SCA3, recent elegant work from Toonen et al, assessed exon-skipping of the CAG repeat containing exon 10 in ATXN3 18 . While this exon-specific approach has the advantage that it would likely prevent deleterious effects of loss of ATXN3 function, whether it is sufficiently robust to effectively silence the disease gene requires further study.…”

“…12,13 Our group and others previously tested nucleotidebased gene silencing strategies as potential diseasemodifying therapy in SCA3, including viral-mediated RNA interference [14][15][16] and exon-skipping approaches. 17,18 Although promising, these prior studies have been limited by various factors including-depending on the studyshort-term treatment, spatially restricted silencing in the central nervous system (CNS), limited reduction of mutant ATXN3 (mutATXN3), or concerns about the extent to which the animal model replicates aspects of the human disease. Here, we set out to perform an efficacy study not limited by these factors, building on our recent proof-ofprinciple study that established widespread ASO delivery and silencing of human mutATXN3 in a mouse model of SCA3.…”

Antisense therapies for movement disorders

“…ATXN3 is a deubiquitinase that controls protein quality involving the ubiquitin proteasome . Different approaches have been made to develop therapeutic ASOs for SCA3, including allele‐specific ASOs antisense to the expanded CAG repeat that sterically block translation, splice switching ASOs to exclude exon 10 encoding the expanded repeat, and MOE gapmer ASOs targeting ATXN3 that lower expression of both the wild‐type and mutant ATXN3 forms . Eight‐week‐old SCA3 (Q84/Q84) that were treated with a MOE gapmer ATXN3 ASO for 4 weeks had significantly reduced ATXN3 expression in multiple brain regions, improved expression of eight genes that are abnormally regulated in SCA3, improved motor phenotype (rotarod, balance beam), and restored Purkinje neuron firing frequency .…”

In vivomolecular signatures of cerebellar pathology in spinocerebellar ataxia type 3

“…SCA3 is one of the polyQ diseases for which clinical trials are forthcoming. ASOs [27][28][29][30] , other RNA interference molecules 31, 32 , and pharmacological agents such as the selective serotonin reuptake inhibitor, citalopram [33][34][35] , showed preclinical efficacy to mitigate molecular, pathological, and phenotypic SCA3-like signs in transgenic mice.…”