生長因子受體結合蛋白：Src homology 2 (Grb2-SH2)在致癌基因Ras的訊息傳遞路徑中扮演重要的角色，其包括：細胞的增生與分化。因此，我們著重在發展Grb2-SH2 domain 的胜?抑制物，希望其能研發成抗癌藥劑。然而，為了保護細胞的完整性，細胞膜的組成就形成了一系列的屏障，但這也限制了胜?抑制物當成抗癌藥劑的功效。在這次研究中，為了增進胜?的細胞穿透性，我們藉由結合細胞穿透胜?至我們的Grb2-SH2 前導胜?抑制物：Fmoc-Glu-Tyr-Aib-Asn-NH2，設計了兩條胜?類似物。藉由應用表面膜漿共振技術的BIAcore 生物感測器來探討合成胜?與Grb2-SH2 domain 之間結合相互作用力。我們也研究每一條胜?在人類乳癌細胞中的生物活性。MDA-MB-231和MCF-7 乳癌細胞分別被處理不同濃度的胜?，而細胞存活率是由細胞增生工具組(3-[4, 5-dimethyl-2-thiazolyl]-2, 5-diphenyl tetrazoliumbromide, MTT)來判定。利用流式細胞儀研究胜?對癌細胞中細胞週期及促細胞凋亡的功效。我們研究結果顯示這兩條合成的胜?對MCF-7 乳癌細胞在50 μM 濃度下就有抗增生的效果。這些體外生物檢測方法的建立對於合成胜?、保健食品或抗癌藥的生物活性研究有很大的幫助。 The growth factor receptor-bound protein-Src homology 2 (Grb2-SH2) plays animportant role in the oncogenic Ras signaling pathway, which involves in cellproliferation and differentiation. Therefore, we focus on developing peptidicinhibitors of the Grb2-SH2 domain as promising anticancer agents. However, cellmembrane constitutes a serious barrier for protecting the integrity of cells, it may limitthe effectiveness of peptidic inhibitors as anti-cancer drugs. In this study, to enhancethe cell permeability of peptides, we designed two peptide analogs by incorporation ofthe cell-penetrating peptide (CPP) into one of our leading peptide inhibitors ofGrb2-SH2, Fmoc-Glu-Tyr-Aib-Asn-NH2. The binding interaction between eachsynthetic peptide and Grb2-SH2 domain was detected by using the surface plasmonresonance (SPR) technology developed with the BIAcore-biosensor. We alsoinvestigated the biological activities of each peptide in human breast cancer cells. TheMDA-MB-231 and MCF-7 breast cancer cells were treated separately with variousconcentrations of peptides; and the cell viability was determined by using the cellproliferation kit (3-[4, 5-dimethyl-2-thiazolyl]-2, 5-diphenyl tetrazolium bromide,MTT). Effects of peptides on the cell cycle progression and apoptosis in cancer cellwere studied by flow cytometry. Our results demonstrated that both of the syntheticpeptides exhibited anti-proliferative effect on MCF-7 cancer cells at the concentrationof 50 μM. Establishment of these in vitro bioassays is very useful for investigation ofthe biological activities of synthetic peptides, functional foods or anti-cancer drugs.
