G₂C₄ targeting antisense oligonucleotides potently mitigate TDP-43 dysfunction in human C9orf72 ALS/FTD induced pluripotent stem cell derived neurons

The G₄C₂ repeat expansion in the C9orf72 gene is the most common genetic cause of Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Many studies suggest that dipeptide repeat proteins produced from this repeat are toxic, yet, the contribution of repeat RNA toxicity is under investigated and even less is known regarding the pathogenicity of antisense repeat RNA. Recently, two clinical trials targeting G₄C₂ (sense) repeat RNA via antisense oligonucleotide failed despite a robust decrease in sense-encoded dipeptide repeat proteins demonstrating target engagement. Here, in this brief report, we show that G₂C₄ antisense, but not G₄C₂ sense, repeat RNA is sufficient to induce TDP-43 dysfunction in induced pluripotent stem cell (iPSC) derived neurons (iPSNs). Unexpectedly, only G₂C₄, but not G₄C₂ sense strand targeting, ASOs mitigate deficits in TDP-43 function in authentic C9orf72 ALS/FTD patient iPSNs. Collectively, our data suggest that the G₂C₄ antisense repeat RNA may be an important therapeutic target and provide insights into a possible explanation for the recent G₄C₂ ASO clinical trial failure.