Correcting tau isoform ratios with a long-acting antisense oligonucleotide alleviates 4R-tauopathy phenotypes

Tau, a microtubule-binding protein linked to tauopathies like Alzheimer's disease and frontotemporal lobar degeneration (FTLD), has 3-repeat (3R) and 4-repeat (4R) isoforms. Accumulation of the 4R-tau is associated with FTLD, progressive supranuclear palsy (PSP), and cortico-basal degeneration (CBD). We previously showed that a loss of fused in sarcoma (FUS) or splicing factor, proline- and glutamine-rich (SFPQ) promoted 4R-tau accumulation, which induced FTLD-like behaviors and neurodegeneration in mice. Here, we developed antisense oligonucleotides (ASOs) modified with 2'-O, 4'-C-ethylene-bridged nucleic acids (ENAs), reducing the 4R-tau/3R-tau ratio while maintaining total tau expression from the MAPT gene. In vitro screening identified EN-06 as the most effective ENA. Intracerebroventricular (ICV) administration of EN-06 corrected the 4R/3R-tau ratio in FUS-silenced humanized tau mice and human iPSC-derived neurons. This treatment ameliorated disease phenotypes, including aberrant behaviors, spine dysmorphology, and neurodegeneration. The half-life of EN-06 after a single ICV administration was approximately 6 months in the brain, with splicing correction effects that persisted for 2 years. The efficacy of EN-06 was higher than that of 2'-O-methoxyethyl (MOE)-modified ASO (MO-06). These findings highlight the potential of ENA-modified ASOs to reduce 4R-tau while preserving total MAPT expression, thus offering a safe and long-acting treatment for 4R-tau-associated tauopathies.