Spinocerebellar ataxia type 3 (SCA3) is caused by an expanded polyglutamine stretch in ataxin-3. While wild-type ataxin-3 has important functions, e.g., as a deubiquitinase, downregulation of mutant ataxin-3 is likely to slow down the course of this fatal disease. We established a screening platform with human neurons of patients and controls derived from induced pluripotent stem cells to test antisense oligonucleotides (ASOs) for their effects on ataxin-3 expression. We identified an ASO that suppressed mutant and wild-type ataxin-3 levels by >90% after a singular treatment. Next, we screened pairs of ASOs designed to selectively target the mutant or the wild-type allele by taking advantage of a SNP (c.987G > C) in ATXN3 that is present in most SCA3 patients. We found ASOmut4 to reduce levels of mutant ataxin-3 by 80% after 10 days while leaving expression of wild-type ataxin-3 largely unaffected. In a long-term study we proved this effect to last for about 4 weeks after a single treatment without signs of neurotoxicity. This study provides proof of principle that allele-specific lowering of poly(Q)-expanded ataxin-3 by selective ASOs is feasible and long lasting, with sparing of wild-type ataxin-3 expression in a human cell culture model that is genetically identical to SCA3 patients.

Antisense oligonucleotides (ASOs) are single-stranded nucleic acid strings that can be used to selectively modify protein synthesis by binding complementary (pre-)mRNA sequences. By specific arrangements of DNA and RNA into a chain of nucleic acids and additional modifications of the backbone, sugar, and base, the specificity and functionality of the designed ASOs can be adjusted. Thereby cellular uptake, toxicity, and nuclease resistance, as well as binding affinity and specificity to its target (pre-)mRNA, can be modified. Several neurodegenerative diseases are caused by autosomal dominant toxic gain-of-function mutations, which lead to toxic protein products driving disease progression. ASOs targeting such mutations—or even more comprehensively, associated variants, such as single nucleotide polymorphisms (SNPs)—promise a selective degradation of the mutant (pre-)mRNA while sparing the wild type allele. By this approach, protein expression from the wild type strand is preserved, and side effects from an unselective knockdown of both alleles can be prevented. This makes allele-specific targeting strategies a focus for future personalized therapies. Here, we provide an overview of current strategies to develop personalized, allele-specific ASO therapies for the treatment of neurodegenerative diseases, such Huntington’s disease (HD) and spinocerebellar ataxia type 3 (SCA3/MJD).