Nature Biomedical Engineering. 2023 Feb 16
Sen Yan, Xiao Zheng, Yingqi Lin, Caijuan Li, Zhaoming Liu, Jiawei Li, Zhuchi Tu, Yu Zhao, Chunhui Huang, Yizhi Chen, Jun Li, Xichen Song, Bofeng Han, Wei Wang, Weien Liang, Liangxue Lai, Xiao-Jiang Li & Shihua Li
Products used in the paper Details Operation
AAV vector packaging The AAV-mini-cmv-spCas9(AAV9-Cas9, AAV-Cas9), AAV-controlgRNA- RFP (AAV9-Ctrl-gRNA-RFP, AAV9-Ctrl-gRNA, Ctrl-gRNA) and AAV9-HTT-gRNA-RFP-20Q (AAV9-HTT-gRNA-20Q, HTT-gRNA-20Q) donor vectors were sent to PackGene Biotech (Guangzhou, China) for viral packaging and production. Request Quote

Research Field: CNS

AAV Serotype: AAV9

Dose: a total of 1–1.5 × 10^12 vg. HD KI Bama pig littermates were used to produce the neonatal pigs. Each of eight 3–7-day-old piglets was injected with AAV-mini-cmv-spCas9 and AAV-Ctrl-gRNA-RFP virus (4 pigs for the control group) or AAV-mini-cmv-spCas9/AAV-HTT-gRNA-RFP-20Q (4 pigs for the treatment group) through the auricular vein with viral particles at 2 × 10^13 genome copies per kg or about 1.8 × 10^13 genome copies per pig.

Routes of Administration: Stereotaxic injection; auricular vein

Animal or cell line strain: 3-month-old piglets


The monogenic nature of Huntington’s disease (HD) and other neurodegenerative diseases caused by the expansion of glutamine-encoding CAG repeats makes them particularly amenable to gene therapy. Here we show the feasibility of replacing expanded CAG repeats in the mutant HTT allele with a normal CAG repeat in genetically engineered pigs mimicking the selective neurodegeneration seen in patients with HD. A single intracranial or intravenous injection of adeno-associated virus encoding for Cas9, a single-guide RNA targeting the HTT gene, and donor DNA containing the normal CAG repeat led to the depletion of mutant HTT in the animals and to substantial reductions in the dysregulated expression and neurotoxicity of mutant HTT and in neurological symptoms. Our findings support the further translational development of virally delivered Cas9-based gene therapies for the treatment of genetic neurodegenerative diseases.

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