併用MK-2206與WZB117於乳癌細胞株之協同機轉探討

乳癌是在全世界女性中最常被診斷的癌症，也是造成女性癌症死亡的第二大主要原因。 PI3K/Akt/mTOR訊息傳導的調控已經被證明在癌細胞之生長、存活和代謝方面上皆扮演重要的角色。MK-2206是一已經進入臨床試驗且具有高效價的異位Akt抑制劑。雖然MK-2206先前在許多癌細胞株中被證明是有效的，但是臨床試驗結果顯示單獨使用MK-2206有治療失敗的情況發生。本篇研究之目的是探討MK-2206在乳癌細胞株中的作用、以及尋找是否有其它藥物具有殺死癌細胞的協同作用。在研究過程中發現MK-2206與WZB117（GLUT1抑制劑）合併使用能夠抑制乳癌細胞的生長。使用Western blot來分析Akt/mTOR訊息傳導通路蛋白表現的改變：包括p-Akt、p-mTOR及其下游p70S6K和4E-BP-1。結果顯示僅抑制Akt可能不足以導致細胞死亡。接續的研究說明MK-2206和WZB117的併用可以使DNA損傷的標記，-H2AX上升。同時，分別參與同源重組（homologous recombination, HR）和非同源末端連接（non-homologous end joining, NHEJ）的DNA修復蛋白Rad51、Ku80在併用時顯著減少。如此說明兩者併用不僅能導致DNA損傷，而且也會阻止DNA修復途徑。之後使用siRNA靜默GLUT1，以檢視GLUT1在合併使用而造成DNA損傷作用所扮演的角色。然而，靜默GLUT1對MK-2206誘導的細胞死亡並沒有顯著影響。於是根據以上結果推測，WZB117造成的DNA損傷作用可能來自非依賴GLUT1的訊息傳導機制。

關鍵字

MK-2206 ； WZB117 ； DNA損傷； DNA修復

並列摘要

Breast cancer is the most commonly diagnosed cancer and is the second leading cause of cancer death in female worldwide. Regulation of the PI3K/Akt/mTOR signaling pathway has been shown to play a crucial role in cancer cell growth, survival, and metabolism. MK-2206 is a potent allosteric Akt inhibitor that has entered clinical trials. Although MK-2206 was proved to be effective in many cancer cell lines, MK-2206 alone in clinical trials has met treatment failure. The purpose of this study was to explore the effect of MK-2206 in breast cancer cell lines and the possible synergistic combination with other agents. In the present study, it was discovered that MK-2206 in combination with WZB117, a GLUT1 inhibitor, could inhibit cell proliferation. Akt/mTOR pathway proteins including p-Akt, p-mTOR and its downstream effectors p70S6K and 4E-BP-1 were analyzed by Western blot. Results showed that inhibition of Akt may not be sufﬁcient to induce cell death. Further investigation revealed that MK-2206 and WZB117 combination could induce -H2AX, a DNA damage marker. Expression of levels of Rad51 and Ku80, participating in homologous recombination (HR) and non-homologous end joining (NHEJ) repair respectively, were significantly decreased in combination treatment. It indicated that MK-2206 and WZB117 combination not only causes DNA damage, but also impairs DNA repair system. Silencing GLUT1 with siRNA was conducted to evaluate the role of GLUT1 participating in DNA damage effect in the combinatorial treatment. However, silencing GLUT1 did not produce significant effect on MK-2206 induced cell death. Accordingly, it is speculated that DNA damage effect caused by WZB117 may result from GLUT1-independent signaling.

並列關鍵字

MK-2206 ； WZB117 ； DNA damage ； DNA repair

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