Targeted Inhibition of the miR-199a/214 Cluster by CRISPR Interference Augments the Tumor Tropism of Human Induced Pluripotent Stem Cell-Derived Neural Stem Cells under Hypoxic Condition.
Targeted Inhibition of the miR-199a/214 Cluster by CRISPR Interference Augments the Tumor Tropism of Human Induced Pluripotent Stem Cell-Derived Neural Stem Cells under Hypoxic Condition.
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DOI：
10.1155/2016/3598542
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发表时间：
2016
影响因子：
4.3
通讯作者：
Zhu D
中科院分区：
文献类型：
--
作者：
Luo Y;Xu X;An X;Sun X;Wang S;Zhu D
人类诱导多能干细胞(HiPSC)提供了一种突破性的方法，有助于克服神经干细胞(NSCs)在靶向癌症基因治疗中应用所带来的伦理和致敏挑战。然而，HiPSC来源的神经干细胞(HIPS-NSCs)的趋瘤能力仍有很大的提高空间。在这里，我们试图通过调控内源性miR-199a/214簇的活性来促进HIPS-NSCs的肿瘤趋向性，miR-199a/214簇参与了低氧刺激的细胞迁移的调节。我们首先开发了一种杆状病毒传递的CRISPR干扰系统(CRISPRi)，它通过RNA引导的催化死亡Cas9(DCas9)立体地阻断miR-199a/214簇启动子中的E-box元件。然后，我们将该CRISPRi系统应用于HIPS-NSCs，并成功地抑制了microRNA基因簇中miR-199a-5p、miR-199a-3p和miR-214的表达。同时，HIF1a、MET、MAPK1等与低氧刺激细胞迁移调控相关的靶基因表达水平上调。进一步的迁移分析表明，靶向抑制miR-199a/214簇显著增强了HIPS-NSCs在体内外的肿瘤趋向性。这些发现表明CRISPRi在基于NSC的肿瘤靶向基因治疗中有一种新的应用。
The human induced pluripotent stem cell (hiPSC) provides a breakthrough approach that helps overcoming ethical and allergenic challenges posed in application of neural stem cells (NSCs) in targeted cancer gene therapy. However, the tumor-tropic capacity of hiPSC-derived NSCs (hiPS-NSCs) still has much room to improve. Here we attempted to promote the tumor tropism of hiPS-NSCs by manipulating the activity of endogenous miR-199a/214 cluster that is involved in regulation of hypoxia-stimulated cell migration. We first developed a baculovirus-delivered CRISPR interference (CRISPRi) system that sterically blocked the E-box element in the promoter of the miR-199a/214 cluster with an RNA-guided catalytically dead Cas9 (dCas9). We then applied this CRISPRi system to hiPS-NSCs and successfully suppressed the expression of miR-199a-5p, miR-199a-3p, and miR-214 in the microRNA gene cluster. Meanwhile, the expression levels of their targets related to regulation of hypoxia-stimulated cell migration, such as HIF1A, MET, and MAPK1, were upregulated. Further migration assays demonstrated that the targeted inhibition of the miR-199a/214 cluster significantly enhanced the tumor tropism of hiPS-NSCs both in vitro and in vivo. These findings suggest a novel application of CRISPRi in NSC-based tumor-targeted gene therapy.
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影响因子：
9
作者：
通讯作者：
--
特纳综合征患者特异性诱导多能干细胞中整个 X 染色体的单亲二体性
影响因子：
33.5
作者：
Luo Y;Zhu D;Du R;Gong Y;Xie C;Xu X;Fan Y;Yu B;Sun X;Chen Y
通讯作者：
Chen Y
影响因子：
4.6
作者：
Chang H;Yi B;Ma R;Zhang X;Zhao H;Xi Y
通讯作者：
Xi Y
影响因子：
14.9
作者：
Ho TT;Zhou N;Huang J;Koirala P;Xu M;Fung R;Wu F;Mo YY
通讯作者：
Mo YY
DOI：
10.1038/mtm.2015.50
发表时间：
2016
期刊：
Molecular therapy. Methods & clinical development
影响因子：
--
作者：
Kwang TW;Zeng X;Wang S
通讯作者：
Wang S