蔡菁華Jinghua-Tsai Chang程聖雯Sheng-Wen Cheng醫學研究所2026-10-299999-12-3120202020-07-13https://handle.ncl.edu.tw/11296/crpq4ahttps://scholars.csmu.edu.tw/handle/123456789/36515我們由Slit2-E15蛋白中鑑定出了8個胺基酸的LT1-3胜肽，並以PEG修飾增加其水溶性。水溶性的LT1-3-PEG可以有效的抑制肺癌細胞的侵襲能力與生長能力，也可以抑制乳癌細胞的生長。我們發現在所有測試的肺癌細胞株中有兩株肺癌細胞株對LT1-3-PEG抑制細胞生長的能力有抗性，H1299和H1975。H1299不表現p53基因，而H1975具有p53R273H和EGFRT790M/L858R的突變。其餘具有野生型或其他突變型p53的細胞株對LT1-3-PEG抑制細胞生長的能力都具有敏感性。而這些細胞中的p53被si-p53干擾之後，LT1-3-PEG就失去抑制細胞生長的能力。當在H1299的細胞中表現野生型的p53時，LT1-3-PEG可以抑制H1299細胞的生長。我們想了解H1975細胞無法被LT1-3-PEG抑制生長是因為p53R273H或EGFRT790M/L858R突變有關。我們使用si-RNA干擾內生性的p53R273H和EGFRT790M/L858R基因，表達野生型p53或EGFR基因，再探討LT1-3-PEG對細胞生長的影響。在H1975中，我們發現LT1-3-PEG抑制只表現野生型p53基因的細胞之生長能力優於同時表現野生型p53和EGFR基因。p53之R273胺基酸對於LT1-3-PEG抑制細胞生長能力扮演重要角色，R273是位於DNA結合的區域以及扮演dominant negative gene的角色，過度表現R273C、R273G、R273H或R273W之p53突變基因無法改變H1299細胞對於LT1-3-PEG抑制細胞生長的抗性。雖然H1299對於LT1-3的生長抑制具有抗性，但使用si-RNA干擾H1299細胞的JNK1基因後，H1299細胞對於LT1-3-PEG抑制細胞生長能力即具有敏感性。同樣的，過度表現R273突變的H1299細胞對於LT1-3-PEG抑制細胞生長有抗性，但是處理si-JNK1也可以使細胞轉變為對LT1-3-PEG具有敏感性。因此，將來可以使用JNK的抑制劑與LT1-3-PEG共同治療不表現p53或有帶有R273熱點突變的p53之肺癌病人。這些結果暗示了LT1-3-PEG抑制細胞生長可能與JNK1和p53之間的交互作用有關。但是，LT1-3-PEG如何透過JNK1和p53抑制細胞生需要進一步的研究。We haveidentified an eight amino acid peptide named LT1-3 peptide from Slit2-E15, which has the ability to inhibit the proliferation and invasion of lung cancer cells. We modified the LT1-3 petide with PEG to increase its solubility, and confirmed that the water-soluble LT1-3-PEG can effectively inhibit the invasion and growth ability of lung cancer cells, and can also inhibit the growth of breast cancer cells. We found that only two lung cancer cell lines tested were resistant to the ability of LT1-3-PEG to inhibit cell growth, H1299 and H1975. H1299 does not express the p53 gene, while H1975 has mutations in p53R273H and EGFRT790M / L858R. The remaining cell lines with wild-type or other mutant p53 are sensitive to LT1-3-PEG-mediated growth inhibition. When p53 in these cells is interfered with by si-p53, LT1-3-PEG loses its ability to inhibit cell growth. When wild-type p53 is expressed in H1299 cells, LT1-3-PEG can inhibit cell growth. To dissect whether p53R273H or EGFRT790M / L858Ris responsible for H1975 cells resistance to LT1-3-PEG-mediated growth inhibition, we used si-RNA to interfere with endogenous p53 R273H and EGFRT790M / L858R genes, and express wild-type p53 and/or EGFR genes and processed LT1-3-PEG. In H1975, we found that LT1-3-PEG inhibited cell growth better when expressing wild-type p53 gene alone than expressing wild-type p53 and EGFR genes simultaneously. R273 codon of p53 appears important to LT1-3-PEG-mediated growth inhibition. R273 is located in the region where DNA binds and plays the role of dominant negative gene. Overexpression of p53 with R273C, R273G, R273H, or R273W mutation in H1299 cells can not make LT1-3-PEG to inhibit cell growth. Although H1299 is resistant to LT1-3-PEG-mediated growth inhibition, knockdown JNK1 expression converts H1299 to LT1-3-PEG sensitive. Similarly, the ectopic expression of R273 mutant p53 in H1299 is LT1-3-PEG resistant, and knockdown the expression of JNK1 also transforms cells into LT1-3-PEG sensitive. Therefore, the co-treatment of LT1-3-PEG and JNK1 inhibitor can be used to treat patients with p53 null or R273 hot spot mutations of p53. These results suggest that LT1 -3-PEG may affect the cell growth through crosstalk between p53 and JNK1. However, how LT1-3-PEG inhibits cell growth through JNK and p53 requires further study.p53Slit2EGFR胜肽生長抑制PeptideGrowth inhibitionthesis