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Publications

Journal Article Ppublish

PubMed: 42905853 DOI: 10.1126/scitranslmed.adz2140

Single-nucleotide polymorphism-directed design of allele-specific antisense oligonucleotides for spinocerebellar ataxia type 3.

Sci Transl Med 18(872):eadz2140

Authors: Jacob Helm, Elena Buena-Atienza, Clemens M Sauter, Linus Wiora, Melanie Kraft, Yvonne Schelling, Jakob Admard, Jeannette Hübener-Schmid, Thomas Klockgether, Matthis Synofzik, Luís Pereira de Almeida, Mafalda Raposo, Paola Giunti, Hector Garcia-Moreno, Magda M Santana, Jennifer Faber, Holger Hengel, Nicolas Casadei, Ludger Schöls, Stefan Hauser

Affiliations

  • German Center for Neurodegenerative Diseases (DZNE), 72076 Tübingen, Germany.
  • Department of Neurology and Hertie Institute for Clinical Brain Research, University of Tübingen, 72076 Tübingen, Germany.
  • Graduate School of Cellular and Molecular Neuroscience, University of Tübingen, 72076 Tübingen, Germany.
  • NGS Competence Center Tübingen (NCCT), 72076 Tübingen, Germany.
  • Institute of Medical Genetics and Applied Genomics, University of Tübingen, 72076 Tübingen, Germany.
  • German Center for Neurodegenerative Diseases (DZNE), 53127 Bonn, Germany.
  • Gene and RNA Therapy Center (GRTC), Faculty of Medicine, University of Tübingen, 72076 Tübingen, Germany.
  • Center for Neuroscience and Cell Biology (CNC-UC), University of Coimbra, 3004-504 Coimbra, Portugal.
  • Center for Innovative Biomedicine and Biotechnology (CIBB), University of Coimbra, 3004-504 Coimbra, Portugal.
  • Gene Therapy Center of Excellence (GeneT), University of Coimbra, 3004-504 Coimbra, Portugal.
  • Faculty of Pharmacy of the University of Coimbra (FFUC), University of Coimbra, 3000-548 Coimbra, Portugal.
  • Instituto de Investigação e Inovação em Saúde (i3S), Universidade do Porto, 4200-125 Porto, Portugal.
  • Faculdade de Ciências e Tecnologia, Universidade dos Açores, 9501-801 Ponta Delgada, Portugal.
  • Ataxia Centre, Department of Clinical and Movement Neurosciences, UCL Queen Square Institute of Neurology, University College London, WC1N 3GB London, UK.
  • Department of Neurogenetics, National Hospital for Neurology and Neurosurgery, University College London Hospitals NHS Foundation Trust, WC1N 3GB London, UK.
  • Institute of Interdisciplinary Research (IIIUC), University of Coimbra, 3030-789 Coimbra, Portugal.
  • Department of Parkinson's Disease, Sleep and Movement Disorders, University Hospital Bonn, 53127 Bonn, Germany.
  • Department of Neuroradiology, University Hospital Bonn, 53127 Bonn, Germany.
  • Center of Rare Diseases, University of Tübingen, 72076 Tübingen, Germany.

Abstract

Spinocerebellar ataxia type 3 (SCA3), also called Machado-Joseph disease, the most common form of autosomal dominant cerebellar ataxia, is caused by a CAG repeat expansion in ATXN3. The mutant protein is prone to aggregation and exerts toxic gain-of-function effects, whereas the wild-type protein has important neuroprotective roles as a deubiquitinase. Current treatment strategies for SCA3 lack specificity for the mutant allele. Identifying genetic variants such as single-nucleotide polymorphisms (SNPs) in linkage with the pathogenic CAG expansion offers a promising avenue to develop allele-specific therapeutic interventions, like antisense oligonucleotides (ASOs). To address this, we conducted allele-specific, targeted, multiplexed nanopore sequencing on 79 patients with SCA3 from Germany, UK, and Portugal. We identified almost 200 polymorphisms and grouped them into three haplotype blocks on the basis of linkage disequilibrium. These SNPs were strongly associated with the CAG expansion. Genotypic modeling of our subcohorts indicated that targeting only two SNPs from different blocks could enable allele-specific treatment of the majority of European patients with SCA3. We evaluated 17 candidate ASOs targeting SNPs from different haplotype blocks. Seven of these ASOs caused allele-preferential reduction of more than 65%. A single ASO application in two SCA3 iPSC-derived cortical neurons (AX1, AX2) achieved up to 90% reduction of mutant ATXN3. Toxicity assessments together with RNA sequencing analyses in AX1 did not indicate evidence of toxicity or widespread transcriptional perturbations. These findings highlight that characterizing heterozygosity rates offers valuable opportunities for selective ASO therapies. Our proof-of-principle targeting with ASOs demonstrates the feasibility of haplotype-informed allele-specific silencing in ATXN3.

MeSH

Oligonucleotides, Antisense, Oligonucleotides, Spinocerebellar Ataxias, Haplotypes, Ataxia, Alleles, Therapeutics, Polymorphism, Single Nucleotide, Cerebellar Ataxia, Machado-Joseph Disease