# Gene editing for Spinocerebellar ataxia type 3 taking advantage of the human ATXN3L paralog as replacement gene

## TL;DR

Researchers explored gene editing strategies for Spinocerebellar ataxia type 3 (SCA3) using a self-inactivating system and a human ATXN3L paralog as a replacement gene.

## Contribution

The study introduces a novel ablate-and-replace gene editing strategy using the ATXN3L paralog for potential SCA3 treatment.

## Key findings

- Ablate experiments achieved 55 ± 18% cerebellar editing of the ATXN3 gene in mice.
- The ablate-and-replace strategy showed similar editing efficiency to ablation alone in SCA3 transgenic mice.
- Immunofluorescence and RT-qPCR analyses supported the potential of the strategy for SCA3 treatment.

## Abstract

Spinocerebellar ataxia type 3 (SCA3) is a rare neurodegenerative disease caused by a CAG expansion of the ataxin-3 gene (ATXN3). SCA3 patients suffer from ataxia, spasticity and dystonia in mid-adulthood, with spinocerebellar dysfunction and degeneration. As a monogenic disease for which only symptomatic treatment is available, ATXN3 is an attractive target for gene editing. We used the KamiCas9, a self-inactivating gene editing system, to explore gene editing strategies suitable for all SCA3 patients. We first tested the deletion of exon 10 or the introduction of a premature stop codon into exon 9. High editing events were observed in vitro, but efficiency was very low in SCA3 transgenic mice. We then evaluated an ablate-and-replace strategy. The ablate experiments resulted in 55 ± 18% cerebellar editing of the ATXN3 gene. A human ATXN3L paralog, expressed in the brains of SCA3 patients, may act as a natural, CRISPR-resistant replacement gene. In a proof-of-principle study, ablate and ablate-and-replace strategies were evaluated in SCA3 transgenic mice. Two months after injection, similar editing efficiencies were obtained in the ablate and ablate-and-replace groups. Immunofluorescence and RT-qPCR analyses of cerebellar markers support the development of this strategy for SCA3 treatment.