title: CRISPR/Cas9-mediated somatic correction of a novel coagulator factor IX gene mutation ameliorates hemophilia in mouse.
abstract: The X-linked genetic bleeding disorder caused by deficiency of coagulator factor IX, hemophilia B, is a disease ideally suited for gene therapy with genome editing technology. Here, we identify a family with hemophilia B carrying a novel mutation, Y371D, in the human F9 gene. The CRISPR/Cas9 system was used to generate distinct genetically modified mouse models and confirmed that the novel Y371D mutation resulted in a more severe hemophilia B phenotype than the previously identified Y371S mutation. To develop therapeutic strategies targeting this mutation, we subsequently compared naked DNA constructs versus adenoviral vectors to deliver Cas9 components targeting the F9 Y371D mutation in adult mice. After treatment, hemophilia B mice receiving naked DNA constructs exhibited correction of over 0.56% of F9 alleles in hepatocytes, which was sufficient to restore hemostasis. In contrast, the adenoviral delivery system resulted in a higher corrective efficiency but no therapeutic effects due to severe hepatic toxicity. Our studies suggest that CRISPR/Cas-mediated in situ genome editing could be a feasible therapeutic strategy for human hereditary diseases, although an efficient and clinically relevant delivery system is required for further clinical studies.
authors.affiliation: Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China.
authors.forename: Yuting
authors.initials: Y
authors.lastname: Guan
authors.affiliation: Hainan Provincial Key Laboratory for Human Reproductive Medicine and Genetic Research, Hainan Reproductive Medical Center, the Affiliated Hospital of Hainan Medical University Hainan Medical University, Haikou, China Department of Obstetrics and Gynecology, Nanfang Hospital, Southern Medical University, Guangzhou, China mayl1990@foxmail.com myliu@bio.ecnu.edu.cn dlli@bio.ecnu.edu.cn.
authors.forename: Yanlin
authors.initials: Y
authors.lastname: Ma
authors.affiliation: Hainan Provincial Key Laboratory for Human Reproductive Medicine and Genetic Research, Hainan Reproductive Medical Center, the Affiliated Hospital of Hainan Medical University Hainan Medical University, Haikou, China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Li
authors.affiliation: Fengxian Hospital affiliated to Southern Medical University, Shanghai, China.
authors.forename: Zhenliang
authors.initials: Z
authors.lastname: Sun
authors.affiliation: Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China.
authors.forename: Lie
authors.initials: L
authors.lastname: Ma
authors.affiliation: Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China.
authors.forename: Lijuan
authors.initials: L
authors.lastname: Wu
authors.affiliation: Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China.
authors.forename: Liren
authors.initials: L
authors.lastname: Wang
authors.affiliation: Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China.
authors.forename: Li
authors.initials: L
authors.lastname: Zeng
authors.affiliation: Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China.
authors.forename: Yanjiao
authors.initials: Y
authors.lastname: Shao
authors.affiliation: Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China.
authors.forename: Yuting
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Hainan Provincial Key Laboratory for Human Reproductive Medicine and Genetic Research, Hainan Reproductive Medical Center, the Affiliated Hospital of Hainan Medical University Hainan Medical University, Haikou, China.
authors.forename: Ning
authors.initials: N
authors.lastname: Ma
authors.affiliation: Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China.
authors.forename: Wenqing
authors.initials: W
authors.lastname: Lu
authors.affiliation: Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China.
authors.forename: Kewen
authors.initials: K
authors.lastname: Hu
authors.affiliation: Bioray Laboratories Inc., Shanghai, China.
authors.forename: Honghui
authors.initials: H
authors.lastname: Han
authors.affiliation: Department of Obstetrics and Gynecology, Nanfang Hospital, Southern Medical University, Guangzhou, China.
authors.forename: Yanhong
authors.initials: Y
authors.lastname: Yu
authors.affiliation: Hainan Provincial Key Laboratory for Human Reproductive Medicine and Genetic Research, Hainan Reproductive Medical Center, the Affiliated Hospital of Hainan Medical University Hainan Medical University, Haikou, China.
authors.forename: Yuanhua
authors.initials: Y
authors.lastname: Huang
authors.affiliation: Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China Department of Molecular and Cellular Medicine, The Institute of Biosciences and Technology, Texas A&M University Health Science Center, Houston, TX, USA mayl1990@foxmail.com myliu@bio.ecnu.edu.cn dlli@bio.ecnu.edu.cn.
authors.forename: Mingyao
authors.initials: M
authors.lastname: Liu
authors.affiliation: Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China mayl1990@foxmail.com myliu@bio.ecnu.edu.cn dlli@bio.ecnu.edu.cn.
authors.forename: Dali
authors.initials: D
authors.lastname: Li
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D064113:CRISPR-Cas Systems; D002648:Child; D004195:Disease Models, Animal; D005164:Factor IX / Q000235:genetics*; D000072669:Gene Editing / Q000379:methods*; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D002836:Hemophilia B / Q000473:pathology* / Q000628:therapy*; D006801:Humans; D008297:Male; D051379:Mice; D020125:Mutation, Missense; D011995:Recombination, Genetic; D016896:Treatment Outcome
chemical_list: D005164:Factor IX
keywords: gene therapy; genome editing; hemophilia B; hemostasis; monogenetic disease