Ajredini, R. | bioRxiv Science

bioRxiv2025

Repeat associated non-AUG translation as a common mechanism for the polyGln ataxias

Determining if repeat associated non-AUG (RAN) proteins contribute to the CAG polyGln-encoding spinocerebellar ataxias (CAG-SCAs) is critical for understanding mechanisms and developing therapies for these diseases. Immunohistochemistry using antibodies against polySer and polyLeu repeats and locus specific C-terminal regions show sense polySer (AGC frame) and antisense polyLeu (CUG frame) RAN proteins accumulate in affected grey and white-matter brain regions, throughout the cerebellum and pons, in SCA1, SCA2, SCA3, SCA6, and SCA7 autopsy brains. Cerebellar white matter regions with prominent polySer and polyLeu but minimal polyGln aggregates show demyelination, white matter loss, and activated microglia. In SCA3 mice, RAN proteins accumulate in an age-dependent manner. In neural cells, polySer and polyLeu RAN proteins are toxic and cause autophagic dysfunction. In cells, the FDA-approved drug metformin decreases RAN protein levels and reduces toxicity. Taken together, these data identify sense and antisense RAN proteins as a common molecular mechanism shared by the CAG-SCAs.

neuroscience↗

bioRxiv2025

Metformin decreases RAN proteins, rescues splicing abnormalities and improves behavioral phenotypes in SCA8 BAC mice

Spinocerebellar ataxia type 8 is one member of a larger group of dominantly inherited, debilitating neurological diseases caused by CTG*CAG expansions for which there are no effective disease-targeting treatments. RAN translation, which was discovered in SCA8, has previously been shown to occur across CAG and CUG expansion transcripts, making treatments that work for SCA8 potentially relevant to a much broader group of diseases, including SCA1, 2, 3, 6, 7, 12, Huntingtons Disease (HD) Fuchs endothelial corneal dystrophy (FECD), and myotonic dystrophy type 1 (DM1). The FDA-approved drug, metformin, has been previously shown to reduce RAN protein levels in cells overexpressing SCA8 CAG repeats. Here we show, using SCA8 BAC transgenic mice, that metformin treatment improves ambulatory performance, including rotarod, DigiGait, and open field measures. At the molecular level, metformin-treated mice show reduced RAN protein levels and improved splicing abnormalities without changing the levels of the expanded RNAs. Metformin-treated mice also show decreased neuroinflammation with reduced levels of astrogliosis and reduced numbers of activated microglia. Taken together, these data provide strong support for testing FDA-approved metformin in clinical trials for SCA8 and potentially the broader group of CAG*CTG repeat expansion disorders. One Sentence SummaryMetformin improves behavior, neuropathological and molecular phenotypes in SCA8 BAC transgenic mice.

neuroscience↗