title: Enhanced plasma factor VIII activity in mice via cysteine mutation using dual vectors.
abstract: Hemophilia A is caused by a genetic mutation in coagulation factor VIII (FVIII) gene and gene therapy is considered to be a promising strategy for its treatment. We recently demonstrated that co-delivery of two vectors expressing M662C mutated heavy and D1828C mutated light chain genes of B-domain-deleted coagulation factor VIII (BDD-FVIII) leads to inter-chain disulfide cross-linking and improved heavy chain secretion in vitro. In this study, co-injection of both M662C and D1828C mutated BDD-FVIII gene expression vectors into mice resulted in increased heavy chain secretion and coagulation activity in plasma in vivo. Approximately (239 ± 56) ng mL(-1) above endogenous levels of transgenic FVIII heavy chain was found in mouse plasma using a chain-specific ELISA. For FVIII coagulation activity, approximately (1.09 ± 0.25) IU mL(-1) above endogenous levels were detected in co-injected transgenic mouse plasma using a chromogenic assay. These data demonstrate that inter-chain disulfide bonds likely increase heavy chain secretion and coagulation activity in the plasma of transgenic mice with an improved efficacy of a dual-vector delivery of BDD-FVIII gene. These findings support our ongoing efforts to develop a gene therapy for hemophilia A treatment using dual-AAV vectors.
authors: Life Science College of Ludong University, Yantai 264025, China. fuxiangmail@163.com
authors: Fuxiang
authors: F
authors: Zhu
authors: Zelong
authors: Z
authors: Liu
authors: Jing
authors: J
authors: Miao
authors: Huige
authors: H
authors: Qu
authors: Xiaoyan
authors: X
authors: Chi
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D003545:Cysteine / Q000235:genetics* / Q000378:metabolism; D003587:Cytomegalovirus / Q000235:genetics; D005169:Factor VIII / Q000235:genetics* / Q000378:metabolism; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors / Q000235:genetics; D006467:Hemophilia A / Q000097:blood / Q000235:genetics / Q000628:therapy; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D009154:Mutation; D010446:Peptide Fragments / Q000235:genetics* / Q000378:metabolism
chemical_list: C451034:B-domain-deleted factor VIII; D010446:Peptide Fragments; D005169:Factor VIII; D003545:Cysteine