Antisense oligonucleotide–mediated MSH3 suppression reduces somatic CAG repeat expansion in Huntington’s disease iPSC–derived striatal neurons

Abstract

Expanded CAG alleles in the huntingtin (<i>HTT</i>) gene that cause the neurodegenerative disorder Huntington's disease (HD) are genetically unstable and continue to expand somatically throughout life, driving HD onset and progression. MSH3, a DNA mismatch repair protein, modifies HD onset and progression by driving this somatic CAG repeat expansion process. <i>MSH3</i> is relatively tolerant of loss-of-function variation in humans, making it a potential therapeutic target. Here, we show that an <i>MSH3</i>-targeting antisense oligonucleotide (ASO) effectively engaged with its RNA target in induced pluripotent stem cell (iPSC)-derived striatal neurons obtained from a patient with HD carrying <i>125 HTT</i> CAG repeats (the 125 CAG iPSC line). ASO treatment led to a dose-dependent reduction of MSH3 and subsequent stalling of CAG repeat expansion in these striatal neurons. Bulk RNA sequencing revealed a safe profile for <i>MSH3</i> reduction, even when reduced by >95%. Maximal knockdown of MSH3 also effectively slowed CAG repeat expansion in striatal neurons with an otherwise accelerated expansion rate, derived from the 125 CAG iPSC line where <i>FAN1</i> was knocked out by CRISPR-Cas9 editing. Last, we created a knock-in mouse model expressing the human <i>MSH3</i> gene and demonstrated effective in vivo reduction in human <i>MSH3</i> after ASO treatment. Our study shows that ASO-mediated MSH3 reduction can prevent <i>HTT</i> CAG repeat expansion in HD 125 CAG iPSC-derived striatal neurons, highlighting the therapeutic potential of this approach.