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Ectopic Expression of FVIII in HPCs and MSCs Derived from hiPSCs with Site-Specific Integration of ITGA2B Promoter-Driven BDDF8 Gene in Hemophilia A.
- Source :
-
International journal of molecular sciences [Int J Mol Sci] 2022 Jan 06; Vol. 23 (2). Date of Electronic Publication: 2022 Jan 06. - Publication Year :
- 2022
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Abstract
- Hemophilia A (HA) is caused by mutations in the coagulation factor VIII (FVIII) gene (F8) . Gene therapy is a hopeful cure for HA; however, FVIII inhibitors formation hinders its clinical application. Given that platelets promote coagulation via locally releasing α-granule, FVIII ectopically expressed in platelets has been attempted, with promising results for HA treatment. The B-domain-deleted F8 ( BDDF8 ), driven by a truncated ITGA2B promoter, was targeted at the ribosomal DNA (rDNA) locus of HA patient-specific induced pluripotent stem cells (HA-iPSCs). The F8 -modified, human induced pluripotent stem cells (2bF8-iPSCs) were differentiated into induced hematopoietic progenitor cells (iHPCs), induced megakaryocytes (iMKs), and mesenchymal stem cells (iMSCs), and the FVIII expression was detected. The ITGA2B promoter-driven BDDF8 was site-specifically integrated into the rDNA locus of HA-iPSCs. The 2bF8-iPSCs were efficiently differentiated into 2bF8-iHPCs, 2bF8-iMKs, and 2bF8-iMSCs. FVIII was 10.31 ng/10 <superscript>6</superscript> cells in lysates of 2bF8-iHPCs, compared to 1.56 ng/10 <superscript>6</superscript> cells in HA-iHPCs, and FVIII was 3.64 ng/10 <superscript>6</superscript> cells in 2bF8-iMSCs lysates, while 1.31 ng/10 <superscript>6</superscript> cells in iMSCs with CMV-driven BDDF8 . Our results demonstrated a high expression of FVIII in iHPCs and iMSCs derived from hiPSCs with site-specific integration of ITGA2B promoter-driven BDDF8 , indicating potential clinical prospects of this platelet-targeted strategy for HA gene therapy.
- Subjects :
- Base Sequence
DNA, Ribosomal genetics
Factor VIII chemistry
Factor VIII metabolism
Gene Targeting
Genetic Loci
Genetic Vectors metabolism
Humans
Integrin alpha2 metabolism
Megakaryocytes metabolism
Protein Domains
Sequence Deletion
Transcription Activator-Like Effector Nucleases metabolism
Ectopic Gene Expression
Factor VIII genetics
Hematopoietic Stem Cells metabolism
Hemophilia A genetics
Induced Pluripotent Stem Cells metabolism
Integrin alpha2 genetics
Mesenchymal Stem Cells metabolism
Promoter Regions, Genetic
Subjects
Details
- Language :
- English
- ISSN :
- 1422-0067
- Volume :
- 23
- Issue :
- 2
- Database :
- MEDLINE
- Journal :
- International journal of molecular sciences
- Publication Type :
- Academic Journal
- Accession number :
- 35054807
- Full Text :
- https://doi.org/10.3390/ijms23020623