Establishment of anti-cancer drug screening platforms for lung cancer: The identification of effective drugs and functional mechanisms

• Main Authors: Yi-Hua Lai, 賴怡樺
• Other Authors: Jeremy J.W.Chen
• Format: Others
• Language: zh-TW
• Published: 2013
• Online Access: http://ndltd.ncl.edu.tw/handle/86hvd5

博士 === 國立中興大學 === 分子生物學研究所 === 101 === High mortality lung cancer is the most frequent cause of cancer deaths worldwide,including Taiwan. Because of cancer cell metastasis, cancer patients have poor prognosis. The process of cancer metastasis, cancer cells via peripheral vascular or lymphatic move to other parts of the division and growth. The process not only regulate tumor suppressor gene or oncogene expression, cancer cells also secrete angiogenic factors to promote angiogenesis and accelerate cancer metastasis. A lot of pharmacologists try to find the answers from the traditional Chinese herb medicines. However, there is no specific and high-throughput way to screen thousands of these “health-care” herbs to unknown anti-cancer pathway. According to our previous studies, we have identified a novel suppressor gene (HLJ1) which might relate to patients’ survival rate in NSCLC and can be used as a potential drug target. The vascular endothelial angiogenic factors (VEGF) plays essential roles in the activation of many downstream signalling pathways, promotion of angiogenesis and cancer metastasis and is expected to be a potent target for cancer therapy. In this study, we establish three platforms of drug screening. The first and second platforms were the HLJ1 and VEGF-targeting drug-screening platforms which analyze the HLJ1 and VEGF promoter activities by reporter gene assay. Utilizing these platforms, we can screen herbal medicines with lung cancer cell growth inhibition and angiogenesis regulation and investigate their mechanisms. Third platform was computer-aided drug design (CADD) method, we used CADD to develope novel c-Src inhibitors. By CADD, it is easy to identify candidate compounds for biological validation, and increase the successful rate of new drug development. Utilizing drug screening platforms of reporter gene, we identified several herbal compounds from a Chinese herbal library with the capacity to enhance HLJ1 promoter activity or inhibit VEGF transcription and thereby inhibited cancer cell migration and invasion. Among the herbal drugs identified the andrographolide most significantly induced HLJ1 expression and suppressed tumorigenesis both in vitro and in vivo. The andrographolide upregulated HLJ1 via JunB activation, which modulates AP-2α binding at the MMP-2 promoter and represses the expression of MMP-2. Microarray transcriptomic analysis was performed to comprehensively depict the andrographolide-regulated signalling pathways. We showed that andrographolide can affect genes that are dominantly involved in the cell cycle, apoptosis and adhesion-related biological signalling, including mitogen-activated protein kinase, focal adhesion and tight junction pathways, indicating the diverse effects of andrographolide on anticancer invasion and proliferation. In addition, we also found that strophanthin could decrease VEGF mRNA expression and inhibit cancer cell invasion, migration, anchorage-independent and -dependent growth and tumor growth. The tube formation assay was revealed that strophanthin can suppress angiogenesis. Furthermore, the expression of VEGF121, VEGF165 and VEGF189 of VEGF isoforms were repressed by strophanthin. The third platform, computational virtual screening of Src inhibitor by docking on Y418 site, is performed to find the candidate drugs which could bind on Src Y418 site. The results showed that several compounds can suppress Src phosphorylation, especially antihelminthic niclosamide. We demonstrated that niclosamide could reduce src phorsphorylation and lung cancer cell viability, and induce apoptosis, suggesting that niclosamide may have the potential for the clinical treatment or prevention of lung cancer progression in humans. Moreover, we also analyzed the effect of structural changes in the niclosamide molecule on its ability. The result showed that one of the niclosamide derivatives (W3312) was more effective than niclosamide in cell viability. In summary, niclosamide can suppress Src activity and related signaling pathway and make great potential for the clinical treatment. In conclusion, the HLJ1-targeting, VEGF-targeting drug-screening platforms and CADD are useful for screening of novel anticancer compounds. Using these platforms, we identified andrographolide, strophanthin and niclosamide potentially as promising new anticancer agents that could suppress tumor growth and invasion in NSCLC.