- 學位論文
沒食子酸, 比咯苯偶氮駢混成衍生物之抗癌機制探討
Evaluation of p53-Dependent and p53-Independent Apoptotic Pathway with the PBDs-Gallic Hybrid Anticancer Agents
摘要
比咯苯偶氮駢(Pyrrolo[2,1-c][1,4]benzodiazepines, PBDs)由鏈黴菌屬分離出的抗癌烷化試劑，抑制細胞內核酸作用。沒食子酸衍生物(Gallic acid derivatives)是常見天然物其特性為抑制微管蛋白動態平衡。為得到更高的抗癌選擇性，取得一系列PBDs-GAs混成試劑，並做抗癌活性與機轉的評估。PBDs-GAs之4G, 5G抑制黑色素瘤細胞A375效果顯著，且對纖維母細胞無顯著傷害。由PBD-GAs引起細胞內DNA損害，並使細胞週期停滯於S phase。其損害修復指標蛋白ATM與ATR表現量增加，促使下游CHK1抑制CDC25進而導致細胞週期停滯。另一方面，ATM所調控的下游蛋白P53也同時被活化，提高Bax表現並抑制Bcl-xL促使粒線體釋出Cytochrome c而啟動Caspase細胞凋亡機制。值得一提的是，P53突變及缺乏的癌細胞RPMI7951、H1299對PBD-GAs短期耐受性較高，但長期而言還是有相當高的細胞抑制與毒殺效果，其成因為溶酶體滲漏，引起下游粒線體凋亡機制。然而A375卻沒有這種現象，我們認為p53在此扮演抑制溶酶體滲漏的角色。最後動物實驗中，4G的確能有效抑制B16小鼠黑色素瘤細胞增長。
並列摘要
Pyrrolo[2,1-c][1,4]benzodiazepines (PBDs) is a potent inhibitor of nucleic acid synthesis. On the other hand, gallic acid (GA) derivatives have the antitumor properties by inhibition of microtubule polymerization. Recently, we have synthesized a series of PBD-GA derivatives based on their antitumor potentials. The compounds 4G and 5G exhibited higher inhibitory activity on various cell lines, and there was no significant cytotoxicity on human dermal fibroblast cells. We have selected human melanoma cell line A375 as the model for further studies. We detected that cell cycle significantly increased the cell percentage at S phase after the compounds treatment. The indicator of DNA damage, ATM/ATR and their downstream (p53/chk1) were activated, and caused cell cycle arrest by the agents. P53 decreased level of BCL-XL induced cytochrome C leakage from mitochondria made caspase dependent apoptosis by the compounds treated. However, we found that the compounds also highly inhibited p53 deficient cell line H1299 via decrease of lysosomal membrane permeabilization (LMP). These effects should induce mitochondria dependent apoptosis. Nevertheless, the agents didn’t induce LMP in A375. Here p53 played the inhibitor of LMP. Taken together, our studies suggest that PBD-GAs possess potential anticancer activity whether p53-dependent or p53-independent in vitro by induced DNA damage and mitochondrial apoptosis pathway. Additionally, 4G also shown to reduce murine melanoma size in a mouse model.