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생화학분자생물학회 (Korean Society for Biochemistry and Molecular Biology)
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Highly efficient genome editing via CRISPR-Cas9 ribonucleoprotein (RNP) delivery in mesenchymal stem cells

  • A Reum Han (Department of Translational Medicine, Asan Medical Institute of Convergence Science and Technology, Asan Medical Center, University of Ulsan College of Medicine) ;
  • Ha Rim Shin (Department of Cell and Genetic Engineering, Asan Medical Institute of Convergence Science and Technology, Asan Medical Center, University of Ulsan College of Medicine) ;
  • Jiyeon Kweon (Department of Cell and Genetic Engineering, Asan Medical Institute of Convergence Science and Technology, Asan Medical Center, University of Ulsan College of Medicine) ;
  • Soo Been Lee (Department of Biochemistry and Molecular Biology, Asan Medical Institute of Convergence Science and Technology, Asan Medical Center, University of Ulsan College of Medicine) ;
  • Sang Eun Lee (Department of Biochemistry and Molecular Biology, Asan Medical Institute of Convergence Science and Technology, Asan Medical Center, University of Ulsan College of Medicine) ;
  • Eun-Young Kim (Department of Biochemistry and Molecular Biology, Asan Medical Institute of Convergence Science and Technology, Asan Medical Center, University of Ulsan College of Medicine) ;
  • Jiyeon Kweon (Department of Cell and Genetic Engineering, Asan Medical Institute of Convergence Science and Technology, Asan Medical Center, University of Ulsan College of Medicine) ;
  • Eun-Ju Chang (Department of Biochemistry and Molecular Biology, Asan Medical Institute of Convergence Science and Technology, Asan Medical Center, University of Ulsan College of Medicine) ;
  • Yongsub Kim (Department of Cell and Genetic Engineering, Asan Medical Institute of Convergence Science and Technology, Asan Medical Center, University of Ulsan College of Medicine) ;
  • Seong Who Kim (Department of Biochemistry and Molecular Biology, Asan Medical Institute of Convergence Science and Technology, Asan Medical Center, University of Ulsan College of Medicine)
  • 투고 : 2023.06.29
  • 심사 : 2023.08.31
  • 발행 : 2024.01.31
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초록

The CRISPR-Cas9 system has significantly advanced regenerative medicine research by enabling genome editing in stem cells. Due to their desirable properties, mesenchymal stem cells (MSCs) have recently emerged as highly promising therapeutic agents, which properties include differentiation ability and cytokine production. While CRISPR-Cas9 technology is applied to develop MSC-based therapeutics, MSCs exhibit inefficient genome editing, and susceptibility to plasmid DNA. In this study, we compared and optimized plasmid DNA and RNP approaches for efficient genome engineering in MSCs. The RNP-mediated approach enabled genome editing with high indel frequency and low cytotoxicity in MSCs. By utilizing Cas9 RNPs, we successfully generated B2M-knockout MSCs, which reduced T-cell differentiation, and improved MSC survival. Furthermore, this approach enhanced the immunomodulatory effect of IFN-r priming. These findings indicate that the RNP-mediated engineering of MSC genomes can achieve high efficiency, and engineered MSCs offer potential as a promising therapeutic strategy.

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과제정보

This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MSIT) (RS-2023-00260462, 2018R1A5A2020732, 2021R1C1C1007162, and 2023R1A2C2004057).

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