Amyotrophic lateral sclerosis (ALS) is an adult-onset motor neuron disease with a mean survival time of three years. The 97% of the cases have TDP-43 nuclear depletion and cytoplasmic aggregation in motor neurons. TDP-43 prevents non-conserved cryptic exon splicing in certain genes, maintaining transcript stability, including ATG4B, which is crucial for autophagosome maturation and Microtubule-associated proteins 1A/1B light chain 3B (LC3B) homeostasis. In ALS mice (G93A), Atg4b depletion worsens survival rates and autophagy function. For the first time, we observed an elevation of LC3ylation in the CNS of both ALS patients and atg4b^−/− mouse spinal cords. Furthermore, LC3ylation modulates the distribution of ATG3 across membrane compartments. Antisense oligonucleotides (ASOs) targeting cryptic exon restore ATG4B mRNA in TARDBP knockdown cells. We further developed multi-target ASOs targeting TDP-43 binding sequences for a broader effect. Importantly, our ASO based in peptide-PMO conjugates show brain distribution post-IV administration, offering a non-invasive ASO-based treatment avenue for neurodegenerative diseases.

肌萎缩侧索硬化症（ALS）是一种成人发病的运动神经元疾病，平均生存时间为三年。 97%的病例在运动神经元中存在TDP-43核耗竭和细胞质聚集。 TDP-43 可防止某些基因中的非保守隐藏外显子剪接，维持转录稳定性，包括ATG4B ，这对于自噬体成熟和微管相关蛋白 1A/1B 轻链 3B (LC3B) 稳态至关重要。在 ALS 小鼠 (G93A) 中， Atg4b耗竭会降低存活率和自噬功能。我们首次观察到 ALS 患者和atg4b ^−/−小鼠脊髓的 CNS 中 LC3 化水平升高。此外，LC3 化调节 ATG3 跨膜区室的分布。靶向隐秘外显子的反义寡核苷酸 (ASO) 可恢复TARDBP敲低细胞中的ATG4B mRNA。我们进一步开发了针对 TDP-43 结合序列的多靶点 ASO，以获得更广泛的效果。重要的是，我们基于肽-PMO 缀合物的 ASO 显示了 IV 给药后的大脑分布，为神经退行性疾病提供了一种基于 ASO 的非侵入性治疗途径。