Isoalantolactone mediates the degradation of BCR-ABL protein in imatinib-resistant CML cells by down-regulating survivin
Isoalantolactone mediates the degradation of BCR-ABL protein in imatinib-resistant CML cells by down-regulating survivin
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DOI：
10.1080/15384101.2023.2209963
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发表时间：
2023-05-20
期刊：
影响因子：
4.3
通讯作者：
Gao，Feng-Hou
中科院分区：
文献类型：
--
作者：
Yin，Shan-Shan;Chen，Chen;Gao，Feng-Hou
异土木香内酯（Iso）是从土木香（Inula helenium L.）中分离得到的一种具有生物活性的内酯类化合物，具有多种药理作用。为探讨异土木香内酯在慢性粒细胞白血病（CML）中的作用及其机制，本研究首先采用CCK 8法研究异土木香内酯对伊马替尼敏感和耐药CML细胞的抗增殖作用。流式细胞仪检测异土木香内酯诱导的细胞凋亡。用慢病毒载体pSIN-3×flag-PURO在KBM 5和KBM 5 T315 I细胞中高效表达Survivin。在KBM 5和KBM 5 T315 I细胞中，使用shRNA敲低Survivin。采用细胞热位移试验（CETSA）检测异土木香内酯与Survivin的相互作用。免疫共沉淀法检测异土木香内酯诱导的survivin泛素表达。采用定量聚合酶链反应（Q-PCR）和蛋白质印迹法检测mRNA和蛋白质水平。异土木香内酯抑制伊马替尼耐药CML细胞增殖并促进其凋亡异土木香内酯虽然抑制BCR-ABL和Survivin蛋白的表达，但不能抑制Survivin和BCR-ABL mRNA的表达。同时，异土木香内酯可以通过增加泛素化来降解Survivin蛋白。结果表明，异土木香内酯诱导的Survivin介导BCR-ABL蛋白的下调。还揭示了异土木香内酯通过半胱天冬酶-3触发BCR-ABL蛋白降解。总之，异土木香内酯通过泛素蛋白酶体途径抑制生存素，并以半胱天冬酶-3依赖性方式介导BCR-ABL下调。这些数据表明，异土木香内酯是一种天然化合物，可用作治疗TKI耐药CML的潜在药物。
Isoalantolactone (Iso) is a bioactive lactone isolated from the root of Inula helenium L, which has been reported to have many pharmacological effects. To investigate the role and mechanism of isoalantolactone in chronic myeloid leukemia (CML), we first investigated isoalantolactone’s anti-proliferative effects on imatinib-sensitive and imatinib-resistant CML cells by CCK8. Flow cytometry was used to detect isoalantolactone-induced cell apoptosis. Survivin was overexpressed in KBM5 and KBM5T315Icells using the lentivirus vector pSIN-3×flag-PURO. In KBM5 and KBM5T315Icells, shRNA was used to knockdown survivin. Cellular Thermal Shift Assay (CETSA) was used to detect the interaction between isoalantolactone and survivin. The ubiquitin of survivin induced by isoalantolactone was detected through immunoprecipitation. Quantitative polymerase-chain reaction (Q-PCR) and western blotting were used to detect the levels of mRNA and protein. Isoalantolactone inhibits the proliferation and promotes apoptosis of imatinib-resistant CML cells. Although isoalantolactone inhibits the proteins of BCR-ABL and survivin, it cannot inhibit survivin and BCR-ABL mRNA levels. Simultaneously, it was shown that isoalantolactone can degrade survivin protein by increasing ubiquitination. It was demonstrated that isoalantolactone-induced survivin mediated downregulation of BCR-ABL protein. It was also revealed that isoalantolactone triggered BCR-ABL protein degradation via caspase-3. Altogether, isoalantolactone inhibits survivin through the ubiquitin proteasome pathway, and mediates BCR-ABL downregulation in a caspase-3 dependent manner. These data suggest that isoalantolactone is a natural compound, which can be used as a potential drug to treat TKI-resistant CML.