Design, Synthesis and Bioactivities Studies of Peptidic Inhibitors of MAPK for the Development of Anticancer Agents

• Main Authors: JHAN,JHAN-RU, 詹湛如
• Other Authors: LUNG,FENG-DI
• Format: Others
• Language: zh-TW
• Published: 2017
• Online Access: http://ndltd.ncl.edu.tw/handle/nz9634

碩士 === 東海大學 === 化學系 === 105 === Cell signaling is an important physiological function of the cell, which causes the cell to respond to external stimulation. Such as cell growth, division, differentiation, and apoptosis. Protein kinase plays an important role in the signaling pathway. Mitogen-activated protein kinase (MAPK) belongs to serine/threonine protein kinases, it can be transduction into cells to the nucleus, activating transcription factors and regulating gene expression, therefore MAPK making an important target for the development of anticancer agents. MAPK is activated by phosphorylation within its activation segment.It was reported that a phosphorylation-sensitive secondary structure could be formed in a 26-amino-acid (LARVADPDHDHTGFLTEYVATRWYRC-NH2, IDA) long synthetic peptide corresponding to the activation segment of Xenopus MAPK, termed IDA MAPK peptide. The unphosphorylated IDA MAPK peptide can specificly inhibit in vitro MAPK activity. Therefore, we selected this sequence as a template to design a series of peptide analogs IDA-1~3and IDA-1-1~3.For the development of peptidic inhibitors of MAPK pathway as anticancer agents. These peptide analogs were synthesized by solid phase peptide synthesis (SPPS) , followed by reverse phase -high performance liquid chromatography (RP-HPLC) purification and matrix assisted laser desorption ionization -time of flight mass spectrometry (MALDI-TOF MS) characterization of synthetic peptides. Characterized peptides were analyzed for their bioactivitions and cell migration by MTT assay、prestoblue assay and wound healing assay in human colon cancer cells (SW480 and HT-29) .The cell permeability of peptides was analyzed by using confocal microscopy.We used western blotting to observe different properties of pMAPK protein kinase. According to the results, IDA-1-3 exhibited greater anti-proliferative effect in the colon cancer cells. The cell permeability of peptides were evaluated by analyzing cells treated with FITC labeled IDA-1 and FAM labeled IDA-1-3 using confocal microscopy, and the results suggested that peptides into HT-29 cancer cells. However, the results of the western blotting did not show the inhibition of protein expression. The results can be used to explore MAPK-mediated signaling. It is a potential lead peptide for further development of anticancer agents.