International Journal of Hematology

DOI: 10.1007/s12185-019-02765-0 Pages: 225-233

CRISPR/Cas9-mediated gene correction in hemophilia B patient-derived iPSCs

1. Kurume University School of Medicine, Division of Hematology and Oncology, Department of Medicine

2. Kyushu University, Center for Advanced Medical Innovation

3. Kumamoto University, Institute of Resource Development and Analysis

4. St. Mary’s Hospital, Center for Hematology and Oncology

Correspondence to:
Koji Nagafuji
Email: knagafuji@med.kurume-u.ac.jp

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Abstract

The clustered regulatory interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) system is an efficient genome-editing tool that holds potential for gene therapy. Here, we report an application of this system for gene repair in hemophilia B (HB) using induced pluripotent stem cells (iPSCs). We prepared targeting plasmids with homology arms containing corrected sequences to repair an in-frame deletion in exon 2 of the factor IX (F9) gene and transfected patient-derived iPSCs with the Cas9 nuclease and a guide RNA expression vector. To validate the expression of corrected F9, we attempted to induce the differentiation of iPSCs toward hepatocyte-like cells (HLCs) in vitro. We successfully repaired a disease-causing mutation in HB in patient-derived iPSCs. The transcription product of corrected F9 was confirmed in HLCs differentiated from gene-corrected iPSCs. Although further research should be undertaken to obtain completely functional hepatocytes with secretion of coagulation factor IX, our study provides a proof-of-principle for HB gene therapy using the CRISPR/Cas9 system.

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