Department of Gene Therapy and Regenerative Medicine, Kagoshima University Graduate School of Medical and Dental Sciences, Kagoshima, Japan.
Center for Innovative Therapy Research and Application, Kagoshima University Graduate School of Medical and Dental Sciences, Kagoshima, Japan.
Stem Cells. 2018 Feb;36(2):230-239. doi: 10.1002/stem.2725. Epub 2017 Nov 8.
The risk of tumor formation poses a challenge for human pluripotent stem cell (hPSC)-based transplantation therapy. Specific and total elimination of tumorigenic hPSCs by suicide genes (SGs) has not been achieved because no methodology currently exists for testing multiple candidate transgene constructs. Here, we present a novel method for efficient generation of tumorigenic cell-targeting lentiviral vectors (TC-LVs) with diverse promoters upstream of a fluorescent protein and SGs. Our two-plasmid system achieved rapid and simultaneous construction of different TC-LVs with different promoters. Ganciclovir (GCV) exerted remarkable cytotoxicity in herpes simplex virus thymidine kinase-transduced hPSCs, and high specificity for undifferentiated cells was achieved using the survivin promoter (TC-LV.Surv). Moreover, GCV treatment completely abolished teratoma formation by TC-LV.Surv-infected hPSCs transplanted into mice, without harmful effects. Thus, TC-LV can efficiently identify the best promoter and SG for specific and complete elimination of tumorigenic hPSCs, facilitating the development of safe regenerative medicine. Stem Cells 2018;36:230-239.
肿瘤形成的风险对基于人类多能干细胞（hPSC）的移植治疗构成了挑战。由于目前尚无测试多个候选转基因构建体的方法，因此尚未实现通过自杀基因（SGs）特异性和完全消除致瘤性 hPSC。在这里，我们提出了一种新的方法，用于在荧光蛋白和 SGs 的上游，通过多种启动子有效地生成针对致瘤性细胞的慢病毒载体（TC-LV）。我们的双质粒系统实现了快速且同时构建具有不同启动子的不同 TC-LV。更昔洛韦（GCV）在单纯疱疹病毒胸苷激酶转导的 hPSC 中表现出明显的细胞毒性，并且使用生存素启动子（TC-LV.Surv）实现了对未分化细胞的高特异性。此外，GCV 处理完全消除了 TC-LV.Surv 感染的 hPSC 移植到小鼠中形成的畸胎瘤，而没有有害影响。因此，TC-LV 可以有效地鉴定出最佳的启动子和 SG，以特异性和完全消除致瘤性 hPSC，从而促进安全的再生医学的发展。干细胞 2018;36:230-239。
