| Summary: | 碩士 === 國立臺灣師範大學 === 人類發展與家庭學系 === 101 === Hepatocellular carcinoma (HCC) is the fifth most common malignancy worldwide. More than 75% cases of HCC occur in the Asia-Pacific region. High mortality of HCC is due to the difficulty in diagnosis and poor prognosis. Chemotherapy is a traditional choice for inoperable HCC, whereas drug resistant limits the therapeutic effect. Thus, there is an urgent need to develop new potential drugs for HCC. Our research group has synthesized a series of compounds for anti-cancer screening using MTT assay. Yao-ram-2-7 is one of them significantly inhibits the growth of HCC Hep 3B cells. Especially, Yao-ram-2-7 displays less cytotoxicity on normal human umbilical vein endothelial cells than the HCC targeted therapy Sorafenib, suggesting Yao-ram-2-7 is safer than Sorafenib. We further show that Yao-ram-2-7 induces apoptosis of Hep 3B cells in a time- and dose-related manner using propidium iodide staining followed by flow cytometry. Increase of cleavage-caspase 3 expression is observed using Western blotting. Yao-ram-2-7 also induces autophagy of Hep 3B cells characterized by the accumulation of acidic vesicular organelles by flow cytometry after staining the cells with acridine orange. Western blot analysis further observed the conversion of
autophagy marker from LC3-I to LC3-II. Compared with Sorafenib, Yao-ram-2-7 induces apoptosis and autophagy at a relatively lower dosage for a shorter period of time. Recently, anticancer and chemopreventive effects of phytochemicals such as curcumin have been suggested. In the present study, combination of Yao-ram-2-7 with curcumin promotes growth inhibition of Hep 3B cells and produces an additivity effect. Cell cycle analysis suggests that the decrease in tumor cell proliferation is due to an increase of G2/M arrest. In contrast, addition of curcumin to Sorafenib displays an antagonism effect, suggesting that patients treated with Sorafenib should avoid food and supplements containing curcumin. In addition, we discover that a GSK-3 inhibitor AR-A014418 and Yao-ram-2-7 have similar biological functions since AR-A014418 alters gene expression of Hep 3B cells similarly to Yao-ram-2-7 by using a bioinformatics database Connectivity Map (CMAP). Western blot and flow cytometric analysis confirm that Yao-ram-2-7 behaves like AR-A014418, inducing autophagy and decreasing protein expression of phospho-GSK-3and total GSK-3 These data demonstrate that query gene expression profiles using CMAP is a useful shortcut to reveal molecular action of a small chemical compound. Taken together, our data suggest chemotherapeutic potential of Yao-ram-2-7 on HCC, and addition of curcumin further promots its chemosensitivity.
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