抗癌胜肽的發展在近幾年越來越受到重視，其對癌細胞的高毒殺性、低抗藥性產生，以及對正常細胞的低副作用，都使其成為研發抗癌藥的新方向。本實驗室從Lactobacillus paracasei ATCC 27092菌株的基因庫中篩選出抗菌胜肽BD21，並證實其具有毒殺人結腸癌細胞SW480的能力。為了更進一步提高毒殺能力，實驗室針對BD21的結構做出修改，增加了帶電性、hydrophobic moment以及α-helix的比例，而得到N3。在本篇研究中，透過CCK8 assay我們知道N3的IC50為6.1 μM，而在caspase抑制劑Z-VAD-fmk和西方墨點法的實驗中，我們發現Z-VAD-fmk並不能抑制N3造成SW480死亡，且SW480內的凋亡指標蛋白PARP，並沒有cleavage form產生，而在培養基中，我們偵測到壞死指標蛋白HMGB1和Cyclophilin A，這證明N3是透過細胞壞死機制造成SW480死亡，而不是凋亡機制。最後透過共軛焦顯微鏡，我們看到N3可以在短時間內打破細胞膜，並進入到細胞中，而細胞型態也隨著時間改變，到最後細胞膜結構完全被破壞，細胞也跟著死亡。

The development of anticancer peptides has received more and more attention in recent years. The high toxicity and low drug resistance of anticancer peptides to cancer cells, and the low side effects of anticancer peptides on normal cells have made them a new direction for the development of anticancer drugs. Our laboratory identified an antimicrobial peptide named BD21 from the gene bank of Lactobacillus paracasei ATCC 27092 and confirmed it has ability to kill human colon adenocarcinoma cell SW480. In order to improve the toxicity, we modified the sequence of BD21 by increasing its electric charge, hydrophobic moment and the ratio of α-helix, and then we got a new peptide N3. In this study, we know that the IC50 of N3 is 6.1μM through the CCK8 assay. By the experiments of caspase inhibitor named Z-VAD-fmk and western bolt, we found that Z-VAD-fmk did not inhibit the death rate of SW480 caused by N3 and the apoptosis marker PARP in SW480 did not produce cleavage form. In the medium, we detected the necrosis marker HMGB1 and Cyclophilin A. These confirmed that N3 induced cell death by necrosis not apoptosis. Finally, through the confocal microscope, we found that N3 could break the cell membrane in a short time and enter the cytoplasm. The cell morphology also changed over time, and finally the cell membrane structure was destructed completely and the cell dead.

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