Preventing autosomal-dominant hearing loss in Bth mice with CRISPR/CasRx-based RNA editing  

作　　者：Ziwen Zheng Guo Li Chong Cui Fang Wang Xiaohan Wang Zhijiao Xu Huiping Guo Yuxin Chen Honghai Tang Daqi Wang Mingqian Huang Zheng-Yi Chen Xingxu Huang Huawei Li Geng-Lin Li Xiaoxiang Hu Yilai Shu 

机构地区：[1]ENT Institute and Department of Otorhinolaryngology,Eye&ENT Hospital,State Key Laboratory of Medical Neurobiology and MOE Frontiers Center for Brain Science,Fudan University,Shanghai 200031,China [2]Institutes of Biomedical Sciences,Fudan University,Shanghai 200032,China [3]NHC Key Laboratory of Hearing Medicine(Fudan University),Shanghai 200032,China [4]State Key Laboratory of Agrobiotechnology,China Agricultural University,Beijing 100193,China [5]College of Biological Sciences,China Agricultural University,Beijing 100193,China [6]School of Life Science and Technology,Southeast University,Nanjing 210096,China [7]Department of Otolaryngology-Head and Neck Surgery,Graduate Program in Speech and Hearing Bioscience and Technology and Program in Neuroscience,Harvard Medical School,Boston,MA 02115,USA [8]Eaton-Peabody Laboratory,Massachusetts Eye and Ear Infirmary,Boston,MA 02114,USA [9]School of Life Science and Technology,ShanghaiTech University,Shanghai 200031,China [10]CAS Center for Excellence in Molecular Cell Science,Shanghai Institute of Biochemistry and Cell Biology,Chinese Academy of Sciences,University of Chinese Academy of Sciences,Shanghai 200031,China [11]Institutes of Brain Science and the Collaborative Innovation Center for Brain Science,Fudan University,Shanghai 200031,China

出　　处：《Signal Transduction and Targeted Therapy》2022年第4期1258-1270,共13页信号转导与靶向治疗（英文）

基　　金：the National Natural Science Foundation of China(82171148,51873107,81770999);National Key Research and Development Program of China(2020YFA0908201);Science and Technology Commission of Shanghai Municipality(21S11905100);National Genetically Modified Organisms Breeding Major Projects of China(2016ZX08009003-006);Clinic Research Plan of SHDC(No.SHDC2020CR4083),“Shuguang Program”supported by Shanghai Education Development Foundation and Shanghai Municipal Education Commission(20SG08);National Research Facility for Phenotypic and Genotypic Analysis of Model Animals(Beijing).

摘　　要：CRISPR/RfxCas13d(CasRx)editing system can specifically and precisely cleave single-strand RNAs,which is a promising treatment for various disorders by downregulation of related gene expression.Here,we tested this RNA-editing approach on Beethoven(Bth)mice,an animal model for human DFNA36 due to a point mutation in Tmc1.We first screened 30 sgRNAs in cell cultures and found that CasRx with sgRNA3 reduced the Tmc1^(Bth)transcript by 90.8%,and the Tmc1 wild type transcript(Tmc1^(+))by 44.3%.We then injected a newly developed AAV vector(AAV-PHP.eB)based CasRx into the inner ears of neonatal Bth mice,and we found that Tmc1^(Bth)was reduced by 70.2%in 2 weeks with few off-target effects in the whole transcriptome.Consistently,we found improved hair cell survival,rescued hair bundle degeneration,and reduced mechanoelectrical transduction current.Importantly,the hearing performance,measured in both ABR and DPOAE thresholds,was improved significantly in all ages over 8 weeks.We,therefore,have validated the CRISPR/CasRx-based RNA editing strategy in treating autosomal-dominant hearing loss,paving way for its further application in many other hereditary diseases in hearing and beyond.

关 键 词：CRISPR/Cas EDITING HEARING 

分 类 号：R764[医药卫生—耳鼻咽喉科]