WRC-213及Mana-Hox對人類前列腺癌細胞之作用機轉探討

The Study of Anticancer Mechanisms of WRC-213 and Mana-Hox in Human Prostate Cancer Cell Lines

蕭哲仁(Che-Jen Hsiao)

指導教授 ： 呂平江 ； 顧記華

國立清華大學/生命科學院/生命科學系/博士(2008年)

摘要

本論文將說明研發藥物WRC-213和Mana-Hox對前列腺癌細胞株(PC-3)之抗癌作用機轉。
第一篇實驗主要是以一系列的實驗，說明anthracenedione類系列衍生物WRC-213引起荷爾蒙不依賴型之前列腺癌細胞株(PC-3)細胞凋亡的機轉。 WRC-213可造成PC-3細胞呈濃度與時間相關性的細胞凋亡現象， WRC-213造成DNA受損並且抑制第二型拓樸異構酶活性，細胞週期停滯在S與G2期，WRC-213引起與粒線體相關的內在性細胞凋亡途徑，造成caspase-3與-9的活化，Mcl-1裂解、Bcl-2表現量下降，而且WRC-213引起survivin表現量減少與caspase-2的活化。另外，WRC-213比臨床抗癌藥物mitoxantrone、etoposide和doxorubicin引起較低的心臟毒性與抗藥性。由實驗證明，WRC-213對治療前列腺癌是一個有潛力的拓樸異構酶作用藥物。
第二篇實驗討論β-carboline類化合物Mana-Hox引起荷爾蒙不依賴型之前列腺癌細胞株(PC-3)細胞凋亡的機轉。從流式細胞儀分析結果顯示Mana-Hox造成細胞週期停滯在G2/M期和造成凋亡細胞增加。從微管聚合實驗和西方墨點法實驗結果顯示，Mana-Hox擾亂微管的聚合並且引起DNA受損作用，Mana-Hox造成cyclin B表現量增加，Cdk1的表現量沒有變化，但是MPM-2表現量增加，而cyclin A及cyclin E 表現量下降。因此推測Mana-Hox引起細胞週期停滯在mitosis期。Mana-Hox引起與粒線體相關的內在性細胞凋亡途徑，也造成Bcl-2蛋白磷酸化，Mcl-1蛋白量減少，PARP與caspase-3活化。而且Mana-Hox在P-glycoprotein相關之抗藥性實驗中呈現較低的抗藥性。綜合上述結果，我們認為Mana-Hox在未來對前列腺癌的化學治療上是極具有潛力的化合物。

關鍵字

前列腺 ； 癌

並列摘要

In the present studies, we elucidate the anticancer mechanism of WRC-213 and Mana-Hox in the androgen-refractory human prostate adenicarcinoma PC-3 cells.
In the first part, the research is to investigate the apoptotic mechanism of WRC-213, an anthracenedione derivative, in human prostate cancer PC-3 cells. WRC-213 induced apoptotic cell death in PC-3 cells in a concentration- and time-dependent manner. WRC-213 induced DNA damage and inhibited activity of topoisomerase II, resulting in cell cycle arrest at S and G2 phase. After the checkpoint arrest, WRC-213 induced mitochondrial-mediated intrinsic apoptotic pathway, including caspase-3/caspase-9 activation, Mcl-1 cleavage and Bcl-2 down-regulation. Survivin degradation and caspase-2 activation also contributed to WRC-213-induced apoptosis. Furthermore, the measurement of cytotoxicity in H9c2 cardiomyocytes and drug resistance in NCI/ADR-RES cells demonstrated that WRC-213 showed much lower cardiotoxicity and P-glycoprotein-related resistance than those of mitoxantrone, etoposide and doxorubicin. In conclusion, WRC-213 is a promising topoisomerase II poison for treatment of androgen-refractory human prostate adenocarcinoma.
In the second part, the research is to investigate the apoptotic mechanism of Mana-Hox, a β-carbolin derivative, in human prostate cancer PC-3 cells. The data from flow cytometric analysis showed that Mana-Hox induced the accumulation of cells at G2/M phase and hypodiploid subG1 phase of the cell cycle. Tubulin assembly and Western blotting assays indicated that Mana-Hox induced tubulin depolymerization and DNA damage. Additionally, Mana-Hox induced up-regulation of cyclin B, down-regulation of cyclin A and cyclin E protein levels, and a marked increase of MPM-2 expression, suggesting the mitotic arrest to Mana-Hox action. Mana-Hox induced the activation of mitochondria-mediated apoptosis pathways and caused phosphorylation of Bcl-2, cleavage of Mcl-1 and PARP, and activation of caspase-3. Moreover, Mana-Hox showed much lower P-glycoprotein-related resistance than taxol, vincristine, etoposide, and doxorubicin. The data suggest that Mana-Hox is a potential compound for the development of prostate cancer chemotherapy.

並列關鍵字

prostate ； cancer

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