基于LC-ESI-MS/MS技术的青葙属抗肿瘤荨麻科类型双环环肽的快速发现及其作用机制研究

Malignant tumor is a disease that severely threatens the lives of human beings. To search for natural leading compounds having antitumor activity with high-efficiency and low-toxicity is one of important strategy for drug development. Urticaceae-type cyclopeptides, mainly found in the seeds of Celosia argentea, are rare bicyclic peptides with novel skeletons. They are reported to remarkably inhibit the polymerization of tubulin. Conventional TLC protosite reaction with ninhydrin reagent showed low sensitivity for detecting cyclopeptides from crude samples, but when combined with sensitive LC-ESI-MS/MS (liquid chromatography-tandem mass spectrometric method using electrospray ionization source) can quickly detect minor cyclopeptides. In the previous work, a new cyclopeptide was firstly isolated by us from Celosia cristata, and showed good inhibitory activities against A549 (human lung cancer cell line) and HepG2 (human hepatoma tumor cell line) for the first time. Based on the previous studies, the present project will carry out the directional isolation of novel Urticaceae-type cyclopeptides from the seeds of C. argentea and C. cristata by LC-ESI-MS/MS and TLC method. Then the chemical constituents and antitumor activities of other compounds will also be studied. Moreover, the inhibitory activities against human tumor cell lines, one normal cell line and the polymerization of tubulin will be screened. Then the cyclopeptide with best activities and the sensitive human tumor cell line will be selected to study their antitumor mechanism on tubulin, cell cycle and apoptosis.

恶性肿瘤是严重威胁人类生命的疾病。寻找高效低毒的天然抗肿瘤活性先导化合物是药物研发的重要战略之一。荨麻科类型环肽是一类少见且骨架新颖的双环环肽，主要发现于青葙属植物青葙子中，被报道具有很强的微管蛋白聚合抑制活性。用常规的茚三酮薄层原位显色法检测粗样中的环肽灵敏度低，若结合LC-ESI-MS/MS技术可以快速发现微量环肽。前期我们首次从同属植物鸡冠花子中分离得到1个新环肽，并首次发现该环肽对A549和HepG2有较好的增殖抑制活性。本项目拟在已有研究基础上，联用茚三酮薄层原位显色法和LC-ESI-MS/MS技术双示踪定向分离青葙子和鸡冠花子中的环肽；同时也分离其他化学成分并进行抗肿瘤活性筛选；开展环肽对多种肿瘤细胞株、正常细胞株增殖抑制和微管蛋白聚合抑制活性筛选；并选择活性最好的环肽和肿瘤敏感株，从微管蛋白、细胞周期、细胞凋亡等方面阐明其抗肿瘤作用机制。

恶性肿瘤是严重威胁人类生命和健康的疾病。荨麻科类型环肽是一类骨架新颖的双环杂环肽，主要发现于青葙子中，曾被报道具有较强的微管聚合抑制活性。本课题对青葙属青葙子和鸡冠花子开展荨麻科类型环肽TLC和LC-MS/MS双示踪导向分离。首先完成青葙子和鸡冠花子化学成分的UPLC-ESI-Q-TOF-MS分析研究。在其指导下，从青葙子中分离鉴定6个环肽，鸡冠花子中分离鉴定5个环肽。对量最大的环肽CS-21开展抗肿瘤活性与作用机制研究，首次发现其对A549细胞有较好的抑制活性，且具有诱导A549细胞凋亡、抑制细胞迁移等作用机制。此外，还从青葙子中分离鉴定30个其他类型化合物，其中5个为新化合物。从鸡冠花子中分离鉴定了25个化合物，其中5个为新化合物。开展青葙子中5个新化合物的肝保护活性筛选与作用机制研究，发现活性新化合物CA-33。本研究为进一步深入开展荨麻科类型环肽抗肿瘤作用机制打下基础。此外，本研究开展的新化合物肝保护活性研究，也为进一步阐明青葙子肝保护的传统药效打下基础。

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