Tanshinone I, a new EZH2 inhibitor restricts normal and malignant hematopoiesis through upregulation of MMP9 and ABCG2.
Tanshinone I, a new EZH2 inhibitor restricts normal and malignant hematopoiesis through upregulation of MMP9 and ABCG2.
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丹参酮 I 是一种新型 EZH2 抑制剂，通过上调 MMP9 和 ABCG2 来限制正常和恶性造血
作者：
Huang Y;Yu SH;Zhen WX;Cheng T;Wang D;Lin JB;Wu YH;Wang YF;Chen Y;Shu LP;Wang Y;Sun XJ;Zhou Y;Yang F;Hsu CH;Xu PF
基本原理：丹参酮是一种来源于丹参的二萜类化合物，是一种特别有前途的用于治疗包括急性髓性白血病（AML）在内的癌症的草药化合物。然而，丹参酮对急性髓细胞白血病的治疗作用及其机制尚不清楚，丹参酮的毒性作用限制了其临床应用。研究方法：本研究利用人白血病细胞系、斑马鱼转基因动物和异种移植模型，研究丹参酮影响正常和异常造血的细胞和分子机制。采用WISH、苏丹黑和邻联茴香胺染色法检测斑马鱼胚胎造血基因的表达。RNA-seq分析显示差异表达基因和富集的基因签名与谭I处理。使用表面等离子体共振（SPR）方法和BIAcore T200（GE Healthcare）测量Tan I的结合亲和力。进行体外甲基转移酶测定以验证Tan I抑制PRC 2复合物的组蛋白酶活性。采用ChIP-qPCR检测EZH 2靶基因H3 K27 me 3水平。结果如下：我们发现丹参酮I（Tanshinone I，Tan I）在体外和异种移植斑马鱼模型中均能抑制人白血病细胞的增殖，同时也能抑制斑马鱼正常和恶性造血。机制研究表明，Tan I通过直接结合EZH 2（一种众所周知的组蛋白H3 K27甲基转移酶）和抑制PRC 2酶活性来调节正常和恶性造血。此外，我们确定MMP 9和ABCG 2是Tan I对EZH 2作用的两个可能的下游基因。结论：总之，这项研究证实了Tan I是一种新的EZH 2抑制剂，并表明MMP 9和ABCG 2是骨髓恶性疾病的两个潜在治疗靶点。
Rationale: Tanshinone, a type of diterpenes derived from salvia miltiorrhiza, is a particularly promising herbal medicine compound for the treatment of cancers including acute myeloid leukemia (AML). However, the therapeutic function and the underlying mechanism of Tanshinone in AML are not clear, and the toxic effect of Tanshinone limits its clinical application. Methods: Our work utilizes human leukemia cell lines, zebrafish transgenics and xenograft models to study the cellular and molecular mechanisms of how Tanshinone affects normal and abnormal hematopoiesis. WISH, Sudan Black and O-Dianisidine Staining were used to determine the expression of hematopoietic genes on zebrafish embryos. RNA-seq analysis showed that differential expression genes and enrichment gene signature with Tan I treatment. The surface plasmon resonance (SPR) method was used with a BIAcore T200 (GE Healthcare) to measure the binding affinities of Tan I. In vitro methyltransferase assay was performed to verify Tan I inhibits the histone enzymatic activity of the PRC2 complex. ChIP-qPCR assay was used to determine the H3K27me3 level of EZH2 target genes. Results: We found that Tanshinone I (Tan I), one of the Tanshinones, can inhibit the proliferation of human leukemia cells in vitro and in the xenograft zebrafish model, as well as the normal and malignant definitive hematopoiesis in zebrafish. Mechanistic studies illustrate that Tan I regulates normal and malignant hematopoiesis through direct binding to EZH2, a well-known histone H3K27 methyltransferase, and inhibiting PRC2 enzymatic activity. Furthermore, we identified MMP9 and ABCG2 as two possible downstream genes of Tan I's effects on EZH2. Conclusions: Together, this study confirmed that Tan I is a novel EZH2 inhibitor and suggested MMP9 and ABCG2 as two potential therapeutic targets for myeloid malignant diseases.