Zu-Jun Que1, Bin Luo2, Chen-Tong Wang3, Fang-Fang Qian4, Yi Jiang5, Yan Li6, Xiang-Hui Han7, He-Gen Li8, Jia-Xiang Liu9, Jian-Hui Tian10. 1. Institute of Traditional Chinese Medicine Oncology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, 200032, China; School of Pharmacy, Shanghai University of Traditional Chinese Medicine, Shanghai, 201203, China. Electronic address: zujunque@shutcm.edu.cn. 2. Institute of Traditional Chinese Medicine Oncology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, 200032, China. Electronic address: luo2014bin@163.com. 3. State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai, 200240, China. Electronic address: 13122310063@163.com. 4. Department of Oncology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, 200032, China. Electronic address: 18883873419@163.com. 5. Department of Oncology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, 200032, China. Electronic address: 877708348@qq.com. 6. Department of Oncology, Shanghai Municipal Hospital of Traditional Chinese Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai, 200071, China. Electronic address: liyan31415@163.com. 7. Institute of Chinese Traditional Surgery, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, 200032, China. Electronic address: hanxianghui1106@163.com. 8. Department of Oncology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, 200032, China. Electronic address: shlaogen@163.com. 9. Department of Oncology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, 200032, China. Electronic address: tjhhawk@163.com. 10. Institute of Traditional Chinese Medicine Oncology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, 200032, China; Department of Oncology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, 200032, China. Electronic address: hawk7150@hotmail.com.
Abstract
ETHNOPHARMACOLOGICAL RELEVANCE: Jinfukang has long been used for the clinical treatment of lung cancer. Previous studies have shown that Jinfukang can induce the apoptosis of circulating tumor cells by intervening ROS-mediated DNA damage pathway. However, whether Jinfukang can inhibit the metastasis of circulating tumor cells and its mechanism are still unclear. AIM OF THE STUDY: To further investigate the mechanism of Jinfukang in anti-metastasis of lung cancer from the perspective of intervention of tumor exosomes. MATERIALS AND METHODS: The invadopodia formation was determined with immunofluorescence. Invasion and migration were detected using the Transwell assay. Ultracentrifugation was used to isolate exosomes. Exosomes were characterized by electron microscopy, nanoparticle tracking analysis and immunoblotting, and the protein profile was evaluated by proteomic analysis. The molecular functions, biological processes and signaling pathway enrichment analysis were performed using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG). Key differentially expressed proteins were verified by Western blot. RESULTS: Jinfukang can inhibit the expression of MMP14, cortactin, Tks5 and the formation of invadopodia of CTC-TJH-01 cells. Furthermore, Jinfukang can significantly inhibit the invasion and migration of CTC-TJH-01 cells. The diameter of exosomes extracted from the CTC-TJH-01 cells treated by Jinfukang was 30-100 nm, and the exosomal markers CD63, CD81 and TSG101 were expressed. We identified 680 deferentially expressed proteins. Gene oncology analysis indicated that exosomes were mostly derived from plasma membrane and mainly involved in protein localization and intracellular signaling. The ingenuity pathway analysis showed that the EGF pathway was significantly inhibited, whereas the GP6 signaling pathway was significantly activated. We also confirmed that Jinfukang inhibited the expression of EGF pathway-related proteins in CTC-TJH-01 cells. Besides, when EGF was used to activate EGF signaling pathway, the inhibition of Jinfukang on CTC cell metastasis was reversed. CONCLUSION: Jinfukang inhibits the metastasis of CTC-TJH-01 cells through the EGF pathway.
ETHNOPHARMACOLOGICAL RELEVANCE: Jinfukang has long been used for the clinical treatment of lung cancer. Previous studies have shown that Jinfukang can induce the apoptosis of circulating tumor cells by intervening ROS-mediated DNA damage pathway. However, whether Jinfukang can inhibit the metastasis of circulating tumor cells and its mechanism are still unclear. AIM OF THE STUDY: To further investigate the mechanism of Jinfukang in anti-metastasis of lung cancer from the perspective of intervention of tumor exosomes. MATERIALS AND METHODS: The invadopodia formation was determined with immunofluorescence. Invasion and migration were detected using the Transwell assay. Ultracentrifugation was used to isolate exosomes. Exosomes were characterized by electron microscopy, nanoparticle tracking analysis and immunoblotting, and the protein profile was evaluated by proteomic analysis. The molecular functions, biological processes and signaling pathway enrichment analysis were performed using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG). Key differentially expressed proteins were verified by Western blot. RESULTS: Jinfukang can inhibit the expression of MMP14, cortactin, Tks5 and the formation of invadopodia of CTC-TJH-01 cells. Furthermore, Jinfukang can significantly inhibit the invasion and migration of CTC-TJH-01 cells. The diameter of exosomes extracted from the CTC-TJH-01 cells treated by Jinfukang was 30-100 nm, and the exosomal markers CD63, CD81 and TSG101 were expressed. We identified 680 deferentially expressed proteins. Gene oncology analysis indicated that exosomes were mostly derived from plasma membrane and mainly involved in protein localization and intracellular signaling. The ingenuity pathway analysis showed that the EGF pathway was significantly inhibited, whereas the GP6 signaling pathway was significantly activated. We also confirmed that Jinfukang inhibited the expression of EGF pathway-related proteins in CTC-TJH-01 cells. Besides, when EGF was used to activate EGF signaling pathway, the inhibition of Jinfukang on CTC cell metastasis was reversed. CONCLUSION: Jinfukang inhibits the metastasis of CTC-TJH-01 cells through the EGF pathway.