Inducible caspase-9 suicide gene under control of endogenous oct4 to safeguard mouse and human pluripotent stem cell therapy

Yang Liu; Yang Yang; Yangyang Suo; Chuan Li; Min Chen; Shuwen Zheng; Hao Li; Chengcheng Tang; Nana Fan; Ting Lan; Jizeng Zhou; Yingying Li; Jiaowei Wang; Huangyao Chen; Qingjian Zou; Liangxue Lai

doi:10.1016/j.omtm.2022.01.014

Inducible caspase-9 suicide gene under control of endogenous oct4 to safeguard mouse and human pluripotent stem cell therapy

Mol Ther Methods Clin Dev. 2022 Feb 1:24:332-341. doi: 10.1016/j.omtm.2022.01.014. eCollection 2022 Mar 10.

Authors

Yang Liu^{1

2

3}, Yang Yang⁴, Yangyang Suo⁵, Chuan Li², Min Chen², Shuwen Zheng², Hao Li³, Chengcheng Tang², Nana Fan³, Ting Lan³, Jizeng Zhou⁴, Yingying Li³, Jiaowei Wang³, Huangyao Chen³, Qingjian Zou², Liangxue Lai^{1

2

3

6

7}

Affiliations

¹ School of Life Sciences, University of Science and Technology of China, Hefei 230026, China.
² Guangdong Provincial Key Laboratory of Large Animal Models for Biomedicine, School of Biotechnology and Health Sciences, Wuyi University, Jiangmen 529020, China.
³ CAS Key Laboratory of Regenerative Biology, Guangdong-Hong Kong Stem Cell and Regenerative Medicine Research Centre, Joint Research Laboratory on Stem Cell and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510530, China.
⁴ School of Biomedical and Pharmaceutical Sciences, Guangdong University of Technology, Guangzhou 510006, China.
⁵ Joint School of Life Science, Guangzhou Institutes of Biomedicine and Health, Chinese Academic and Sciences, Guangzhou Medical University, Guangzhou 511495, China.
⁶ Bioland Laboratory (Guangzhou Regenerative Medicine and Health Guangdong Laboratory), Guangzhou 510005, China.
⁷ Research Unit of Generation of Large Animal Disease Models, Chinese Academy of Medical Sciences (2019RU015), Guangzhou 510530, China.

Abstract

Pluripotent stem cells (PSCs) are promising in regenerative medicine. A major challenge of PSC therapy is the risk of teratoma formation because of the contamination of undifferentiated stem cells. Constitutive promoters or endogenous SOX2 promoters have been used to drive inducible caspase-9 (iCasp9) gene expression but cannot specifically eradicate undifferentiated PSCs. Here, we inserted iCasp9 gene into the endogenous OCT4 locus of human and mouse PSCs without affecting their pluripotency. A chemical inducer of dimerization (CID), AP1903, induced iCasp9 activation, which led to the apoptosis of specific undifferentiated PSCs in vitro and in vivo. Differentiated cell lineages survived because of the silence of the endogenous OCT4 gene. Human and mouse PSCs were controllable when CID was administrated within 2 weeks after PSC injection in immunodeficient mice. However, an interval longer than 2 weeks caused teratoma formation and mouse death because a mass of somatic cells already differentiated from the PSCs. In conclusion, we have developed a specific and efficient PSC suicide system that will be of value in the clinical applications of PSC-based therapy.

Keywords: OCT4 gene; apoptosis; inducible caspase-9 (iCasp9) gene; pluripotent stem cells (PSCs); teratoma formation.