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Mol. Cells

Published online March 21, 2022

© The Korean Society for Molecular and Cellular Biology

Improving the Safety of Mesenchymal Stem Cell-Based Ex Vivo Therapy Using Herpes Simplex Virus Thymidine Kinase

Narayan Bashyal1,2,5 , Tae-Young Lee3,5 , Da-Young Chang3 , Jin-Hwa Jung1 , Min Gyeong Kim1,2 , Rakshya Acharya1 , Sung-Soo Kim1,2 , Il-Hoan Oh4,* , and Haeyoung Suh-Kim1,2,3,*

1Department of Anatomy, Ajou University School of Medicine, Suwon 16499, Korea, 2Department of Biomedical Sciences, Graduate School, Ajou University School of Medicine, Suwon 16499, Korea, 3Research Center, Cell&Brain Co., Ltd., Jeonju 54871, Korea, 4Department of Medical Lifescience, The Catholic University of Korea, College of Medicine, Seoul 06591, Korea, 5These authors contributed equally to this work

Correspondence to : iho@catholic.ac.kr(IHO); hysuh@ajou.ac.kr (HSK)

Received: November 8, 2021; Revised: November 26, 2021; Accepted: December 16, 2021

This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike 3.0 Unported License. To view a copy of this license, visit http://creativecommons.org/licenses/by-nc-sa/3.0/.

Abstract

Human mesenchymal stem cells (MSCs) are multipotent stem cells that have been intensively studied as therapeutic tools for a variety of disorders. To enhance the efficacy of MSCs, therapeutic genes are introduced using retroviral and lentiviral vectors. However, serious adverse events (SAEs) such as tumorigenesis can be induced by insertional mutagenesis. We generated lentiviral vectors encoding the wild-type herpes simplex virus thymidine kinase (HSV-TK) gene and a gene containing a point mutation that results in an alanine to histidine substitution at residue 168 (TK(A168H)) and transduced expression in MSCs (MSC-TK and MSC-TK(A168H)). Transduction of lentiviral vectors encoding the TK(A168H) mutant did not alter the proliferation capacity, mesodermal differentiation potential, or surface antigenicity of MSCs. The MSC-TK(A168H) cells were genetically stable, as shown by karyotyping. MSC-TK(A168H) responded to ganciclovir (GCV) with an half maximal inhibitory concentration (IC50) value 10-fold less than that of MSC-TK. Because MSC-TK(A168H) cells were found to be non-tumorigenic, a U87-TK(A168H) subcutaneous tumor was used as a SAE-like condition and we evaluated the effect of valganciclovir (vGCV), an oral prodrug for GCV. U87-TK(A168H) tumors were more efficiently ablated by 200 mg/kg vGCV than U87-TK tumors. These results indicate that MSC-TK(A168H) cells appear to be pre-clinically safe for therapeutic use. We propose that genetic modification with HSV-TK(A168H) makes allogeneic MSC-based ex vivo therapy safer by eliminating transplanted cells during SAEs such as uncontrolled cell proliferation.

Keywords herpes simplex virus-thymidine kinase, lentiviral vector, mesenchymal stem cells, safety switch, stemness

Article

On-line First

Mol. Cells

Published online March 21, 2022

Copyright © The Korean Society for Molecular and Cellular Biology.

Improving the Safety of Mesenchymal Stem Cell-Based Ex Vivo Therapy Using Herpes Simplex Virus Thymidine Kinase

Narayan Bashyal1,2,5 , Tae-Young Lee3,5 , Da-Young Chang3 , Jin-Hwa Jung1 , Min Gyeong Kim1,2 , Rakshya Acharya1 , Sung-Soo Kim1,2 , Il-Hoan Oh4,* , and Haeyoung Suh-Kim1,2,3,*

1Department of Anatomy, Ajou University School of Medicine, Suwon 16499, Korea, 2Department of Biomedical Sciences, Graduate School, Ajou University School of Medicine, Suwon 16499, Korea, 3Research Center, Cell&Brain Co., Ltd., Jeonju 54871, Korea, 4Department of Medical Lifescience, The Catholic University of Korea, College of Medicine, Seoul 06591, Korea, 5These authors contributed equally to this work

Correspondence to:iho@catholic.ac.kr(IHO); hysuh@ajou.ac.kr (HSK)

Received: November 8, 2021; Revised: November 26, 2021; Accepted: December 16, 2021

This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial-ShareAlike 3.0 Unported License. To view a copy of this license, visit http://creativecommons.org/licenses/by-nc-sa/3.0/.

Abstract

Human mesenchymal stem cells (MSCs) are multipotent stem cells that have been intensively studied as therapeutic tools for a variety of disorders. To enhance the efficacy of MSCs, therapeutic genes are introduced using retroviral and lentiviral vectors. However, serious adverse events (SAEs) such as tumorigenesis can be induced by insertional mutagenesis. We generated lentiviral vectors encoding the wild-type herpes simplex virus thymidine kinase (HSV-TK) gene and a gene containing a point mutation that results in an alanine to histidine substitution at residue 168 (TK(A168H)) and transduced expression in MSCs (MSC-TK and MSC-TK(A168H)). Transduction of lentiviral vectors encoding the TK(A168H) mutant did not alter the proliferation capacity, mesodermal differentiation potential, or surface antigenicity of MSCs. The MSC-TK(A168H) cells were genetically stable, as shown by karyotyping. MSC-TK(A168H) responded to ganciclovir (GCV) with an half maximal inhibitory concentration (IC50) value 10-fold less than that of MSC-TK. Because MSC-TK(A168H) cells were found to be non-tumorigenic, a U87-TK(A168H) subcutaneous tumor was used as a SAE-like condition and we evaluated the effect of valganciclovir (vGCV), an oral prodrug for GCV. U87-TK(A168H) tumors were more efficiently ablated by 200 mg/kg vGCV than U87-TK tumors. These results indicate that MSC-TK(A168H) cells appear to be pre-clinically safe for therapeutic use. We propose that genetic modification with HSV-TK(A168H) makes allogeneic MSC-based ex vivo therapy safer by eliminating transplanted cells during SAEs such as uncontrolled cell proliferation.

Keywords: herpes simplex virus-thymidine kinase, lentiviral vector, mesenchymal stem cells, safety switch, stemness

Mol. Cells
Jun 30, 2022 Vol.45 No.6, pp. 353~434
COVER PICTURE
ERα is modified by UFM1 and this modification (ufmylation) plays a crucial role in promoting the stability of ERα and breast cancer development. However, when ERα is deufmylated and then ubiquitinated, it disappears by proteasome-mediated degradation (Yoo et al., pp. 425-434).

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