Allele-specific targeting of mutant ataxin-3 by antisense oligonucleotides in SCA3-iPSC-derived neurons

Abstract Spinocerebellar ataxia type 3 (SCA3) is caused by an expanded polyglutamine stretch in ataxin-3. While wildtype ataxin-3 has important function 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 its effects on ataxin-3 expression. We identified an ASO that suppressed mutant and wildtype ataxin-3 levels by >90% after a singular treatment. Next, we screened pairs of ASOs designed to selectively target the mutant or the wildtype 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 wildtype ataxin-3 largely unaffected. In a long-term study we proved this effect to last for about 4 weeks after a singular treatment without signs of neurotoxicity. This study provides proof of principle that allele-specific lowering of polyQ-expanded ataxin-3 by selective ASOs is feasible and long lasting with sparing of wildtype ataxin-3 expression in a human cell culture model that is genetically identical to SCA3 patients.