Identification of compound CA-5f as a novel late-stage autophagy inhibitor with potent anti-tumor effect against non-small cell lung cancer
Identification of compound CA-5f as a novel late-stage autophagy inhibitor with potent anti-tumor effect against non-small cell lung cancer
复制标题
化合物CA-5f作为新型新型自噬抑制剂的鉴定，对非小细胞肺癌具有强效抗肿瘤作用
DOI：
10.1080/15548627.2018.1511503
复制
发表时间：
2019-03-04
期刊：
影响因子：
13.3
通讯作者：
Sun, Gangchun
中科院分区：
文献类型：
--
作者：
Zhang, Lu;Qiang, PengFei;Sun, Gangchun
目前，人们特别关注自噬在包括癌症在内的各种人类疾病中的影响。自噬小分子调节剂的发现及其作为抗癌治疗药物的潜在应用将具有重要意义。为此，我们对实验室已合成的一系列姜黄素类似物进行了筛选。在这些化合物中，(3E，5E)-3-(3，4-dimethoxybenzylidene)-5-[(1H-indol-3-yl)methylene]-1-methylpiperidin-4-one(CA-5f)通过抑制自噬小体-溶酶体融合而被鉴定为一种有效的晚期巨噬/自噬抑制因子。我们发现CA-5f既不影响水解酶的功能，也不影响溶酶体的数量。基于相对和绝对定量等压标记(ITRAQ)的蛋白质组筛选结合生物信息学分析表明，CA-5f处理人脐静脉内皮细胞(HUVECs)1h后，细胞骨架蛋白和膜运输蛋白的水平受到抑制。随后的研究表明，CA-5f对A549非小细胞肺癌(NSCLC)细胞具有较强的细胞毒性，但对正常人脐静脉内皮细胞(HUVECs)的细胞毒性较低，其机制是增加线粒体衍生的活性氧(ROS)的产生。此外，CA-5f作为单一药物有效地抑制了A549肺癌移植瘤的生长，具有良好的体内耐受性。Western印迹、免疫荧光和TdT介导的dUTP缺口末端标记(TUNEL)检测结果表明，CA-5f抑制自噬通量，诱导细胞凋亡，而不影响体内CTSB(组织蛋白酶B)和CTSD(组织蛋白酶D)水平，这与体外实验结果一致。综上所述，这些结果表明CA-5f是一种新的晚期自噬抑制剂，具有潜在的临床应用价值。
Currently, particular focus is placed on the implication of autophagy in a variety of human diseases, including cancer. Discovery of small-molecule modulators of autophagy as well as their potential use as anti-cancer therapeutic agents would be of great significance. To this end, a series of curcumin analogs previously synthesized in our laboratory were screened. Among these compounds, (3E,5E)-3-(3,4-dimethoxybenzylidene)-5-[(1H-indol-3-yl)methylene]-1-methylpiperidin-4-one (CA-5f) was identified as a potent late-stage macroautophagy/autophagy inhibitor via inhibiting autophagosome-lysosome fusion. We found that CA-5f neither impaired the hydrolytic function nor the quantity of lysosomes. Use of an isobaric tag for relative and absolute quantitation (iTRAQ)-based proteomic screen in combination with bioinformatics analysis suggested that treatment of human umbilical vein endothelial cells (HUVECs) with CA-5f for 1 h suppressed the levels of cytoskeletal proteins and membrane traffic proteins. Subsequent studies showed that CA-5f exhibited strong cytotoxicity against A549 non-small cell lung cancer (NSCLC) cells, but low cytotoxicity to normal human umbilical vein endothelial cells (HUVECs), by increasing mitochondrial-derived reactive oxygen species (ROS) production. Moreover, CA-5f effectively suppressed the growth of A549 lung cancer xenograft as a single agent with an excellent tolerance in vivo. Results from western blot, immunofluorescence, and TdT-mediated dUTP nick end labeling (TUNEL) assays showed that CA-5f inhibited autophagic flux, induced apoptosis, and did not affect the level of CTSB (cathepsin B) and CTSD (cathepsin D) in vivo, which were consistent with the in vitro data. Collectively, these results demonstrated that CA-5f is a novel late-stage autophagy inhibitor with potential clinical application for NSCLC therapy.