title: Targeting of the human F8 at the multicopy rDNA locus in Hemophilia A patient-derived iPSCs using TALENickases.
abstract: Hemophilia A (HA) is a monogenic disease due to lack of the clotting factor VIII (FVIII). This deficiency may lead to spontaneous joint hemorrhages or life-threatening bleeding but there is no cure for HA until very recently. In this study, we derived induced pluripotent stem cells (iPSCs) from patients with severe HA and used transcription activator-like effector nickases (TALENickases) to target the factor VIII gene (F8) at the multicopy ribosomal DNA (rDNA) locus in HA-iPSCs, aiming to rescue the shortage of FVIII protein. The results revealed that more than one copy of the exogenous F8 could be integrated into the rDNA locus. Importantly, we detected exogenous F8 mRNA and FVIII protein in targeted HA-iPSCs. After they were differentiated into endothelial cells (ECs), the exogenous FVIII protein was still detectable. Thus, it is showed that the multicopy rDNA locus could be utilized as an effective target site in patient-derived iPSCs for gene therapy. This strategy provides a novel iPSCs-based therapeutic option for HA and other monogenic diseases.
authors.affiliation: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China.
authors.forename: Jialun
authors.initials: J
authors.lastname: Pang
authors.affiliation: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China.
authors.forename: Yong
authors.initials: Y
authors.lastname: Wu
authors.affiliation: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China.
authors.forename: Zhuo
authors.initials: Z
authors.lastname: Li
authors.affiliation: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China.
authors.forename: Zhiqing
authors.initials: Z
authors.lastname: Hu
authors.affiliation: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China.
authors.forename: Xiaolin
authors.initials: X
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China.
authors.forename: Xuyun
authors.initials: X
authors.lastname: Hu
authors.affiliation: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China.
authors.forename: Xiaoyan
authors.initials: X
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China.
authors.forename: Xionghao
authors.initials: X
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China.
authors.forename: Miaojin
authors.initials: M
authors.lastname: Zhou
authors.affiliation: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China.
authors.forename: Bo
authors.initials: B
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China.
authors.forename: Yanchi
authors.initials: Y
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China.
authors.forename: Mai
authors.initials: M
authors.lastname: Feng
authors.affiliation: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha, Hunan, China. Electronic address: liangdesheng@sklmg.edu.cn.
authors.forename: Desheng
authors.initials: D
authors.lastname: Liang
mesh_terms: D000818:Animals; D018920:Coculture Techniques; D004275:DNA, Ribosomal / Q000235:genetics; D003850:Deoxyribonuclease I; D005169:Factor VIII / Q000235:genetics*; D015870:Gene Expression; D018390:Gene Targeting / Q000379:methods*; D015316:Genetic Therapy / Q000379:methods; D006467:Hemophilia A / Q000097:blood / Q000235:genetics* / Q000628:therapy*; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000378:metabolism; D008297:Male; D051379:Mice; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014556:Urine / Q000166:cytology
chemical_list: D004275:DNA, Ribosomal; D012333:RNA, Messenger; C078147:F8 protein, human; D005169:Factor VIII; D003850:Deoxyribonuclease I
keywords: Hemophilia A; Multicopy; Ribosomal DNA; TALENickases; iPSCs