Abstract
Neuronal intranuclear inclusion disease (NIID) is an adult-onset neurodegenerative disease caused by expanded GGC repeats in the 5’ untranslated region of the human-specific NOTCH2NLC gene. The high sequence similarity between NOTCH2NLC and its paralogs poses a significant challenge for precise gene editing. Here, we develop a CRISPR/spCas9-based gene-editing strategy that precisely excises the expanded GGC repeats in NOTCH2NLC without detectable off-target effects on the highly homologous NOTCH2/NOTCH2NL family genes (<2% sequence divergence at this locus). The efficacy, specificity and safety of this approach are rigorously validated across multiple experimental models, including human cell lines, NIID iPSCs, and our previously established transgenic NIID mouse model. Our results demonstrate that precise excision of the expanded GGC repeats effectively alleviates NIID-related neuropathological, molecular and behavioral abnormalities. This study establishes the proof of concept for genome editing as a therapeutic strategy for NIID and other related repeat expansion disorders.
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Data availability
The raw RNA-seq data from mice have been deposited in the Gene Expression Omnibus under accession code GSE295763. The raw RNA-seq data from human NPCs and the raw WGS data from human iPSCs have been deposited in the Genome Sequence Archive for Human under accession code HRA011636 and HRA014016, respectively. The deposition and sharing of the raw data have been approved by the Human Genetics Resource Office in China (registration number: 2025BAT00839). All data needed to evaluate the conclusions in the paper are present in the paper and/or the Supplementary data files. Source data are provided with this paper.
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Acknowledgements
The authors would like to thank Peng Jin and Yujing Li from School of Medicine, Emory University; Hao Wu from School of Computer Science and Control Engineering, Shenzhen University of Advanced Technology; Su Yang from Guangdong-Hongkong-Macau Institute of CNS Regeneration, Jinan University; Ying Cheng from Institute of biomedical research, Yunnan University for providing valuable advice to the study. This study was supported by the National Natural Science Foundation of China (82394421 and 82394420 to B.T., 82394422 and 82371874 to X.-J.L., 82271902 and U24A6013 to S.L., 32071037 to Q.L., 82171843 to Y.P., 82101946 to N.X., 82171256 to Q.S.), the National Key R&D Program of China (2021YFA0805200 to H.J.), and the Natural Science Foundation of Hunan Province (2023JJ10097 to Q.L., 2025JJ20079 to Y.P., 2022JJ40832 to N.X.).
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Q.L., Y.P., S.L., and X.-J.L. designed the study, supervised the study, and revised the manuscript; N.X., Y.P., and Q.L. performed experiments, analyzed data, and wrote the manuscript; H.T., Y.L., Y.J., Z.W., J.W., W.Z., X.W., X.S., S.Y., P.Y., Q.S., C.Q., and Y.T. provided important technical assistance to the study; L.S., H.J., D.L., and B.T. provided insightful advice to the study.
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Xie, N., Pan, Y., Tong, H. et al. Precise excision of expanded GGC repeats in NOTCH2NLC via CRISPR/Cas9 for treating neuronal intranuclear inclusion disease. Nat Commun (2026). https://doi.org/10.1038/s41467-026-68385-5
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DOI: https://doi.org/10.1038/s41467-026-68385-5
