三联吡啶衍生物的合成、DNAs相互作用及生物活性研究

发布时间：2018-05-14 00:34

  本文选题：四链体DNAs + 拓扑异构酶　； 参考：《山西大学》2014年硕士论文

【摘要】：本论文设计合成了两种新颖的含有香豆素基团和苯并噻唑基团的三联吡啶衍生物(1)和三联吡啶衍生物(2),并通过多种实验手段探究了目标化合物1与人端粒(h-telo、i-motif)和原癌基因启动子(c-myc、c-kit2)四链体DNAs的相互作用,同时测试了目标化合物1对端粒酶、拓扑异构酶I、癌细胞增殖及其细胞周期进程的影响。实验结果如下：1.在FRET-melting实验中,当目标化合物1的浓度增加到3 rtM时,Fh-teloT、 Fc-mycT和Fc-kit2T的熔点温度变化(△Tm)值达到了15.0、13.6和9.3。C,这表明目标化合物1能够稳定h-telo、c-myc和c-kit2 G-四链体DNAs,且对h-tclo G-四链体DNA的稳定效果好。PCR-Stop实验进一步证明了目标化合物1能够在104 M浓度较好地抑制三种G-四链体DNAs的PCR扩增产物。UV-melting实验证实目标化合物1也能够稳定人端粒i-motif DNA结构,但是不能稳定ct-DNA。2.竞争FRET-melting实验结果表明,在25倍过量的双螺旋ds26存在时,Fc-mycT和Fc-kit2T的△%值没有明显的变化,说明目标化合物1优先与四链体结合；而在50倍过量的双螺旋ds26存在时,Fc-kit2T的△Tm值减小幅度较大,说明当双螺旋DNA的浓度增大时,目标化合物1对Fc-kit2T的选择性降低；在10倍过量双螺旋ds26存在下,FA-teloT的ATm值从9.6℃减小到4.6℃,说明相对于双螺旋ds26,目标化合物1对Fh-teloT的选择性较差。FID实验结果也显示,目标化合物1不能有效置换出与三个四链体DNAs结合的TO,可能是由于二者在DNA上的作用位点不同。3.紫外可见吸收滴定实验结果表明目标化合物1能够与h-telo、c-myc、c-kit2和i-motif四链体DNAs以及双螺旋ct-DNA相互作用且结合常数Kb约为105 M-1。CD实验结果表明：在无阳离子存在下,目标化合物1不能诱导h-telo DNA形成四链体结构；在100 mM K+条件下,目标化合物1能够诱导杂交构象的h-teloG-四链体DNA逐渐地形成反平行构象。且目标化合物1只对c-myc和c-kit2G-四链体DNAs的构象稍有扰动,而且几乎不影响i-motif四链体DNA的构象。4. TRAP实验证实：目标化合物1能够在10-5M浓度有效地抑制人端粒酶的活性,表明目标化合物1是一种潜在的端粒酶活性抑制剂。5.凝胶迁移率实验结果证实：目标化合物1没有使质粒pBR322 DNA发生切割。琼脂糖凝胶电泳实验结果显示,目标化合物1能够以浓度依赖型方式显著地抑制Topo I调控的质粒pBR322 DNA的松弛。且对三联吡啶衍生物而言,香豆素基团的引入和Pt(Ⅱ)的配位提高了它的Topo I抑制活性。6.MTT实验结果证实：目标化合物1能够以浓度依赖型方式抑制HepG2和MCF7癌细胞的增殖,其IC50值分别为3.66和1.54 μM。细胞周期测试结果表明,目标化合物1将HepG2癌细胞的细胞周期进程阻滞于S期,同时将MCF7癌细胞的细胞周期进程阻滞于G0/G1和S期。
[Abstract]:In this paper, we have designed and synthesized two novel tripyridine derivatives containing coumarin group and benzothiazole group. The interaction of DNAs in c-myctropho-c-kit2) of oncogene promoter. The effects of target compound 1 on telomerase, topoisomerase I, cancer cell proliferation and cell cycle progression were also tested. The results of the experiment are as follows: 1. In the FRET-melting experiment, When the concentration of target compound 1 was increased to 3 rtM, the melting point temperature (Tm) of Fh-teloT, Fc-mycT and Fc-kit2T reached 15.0 ~ 13.6 and 9.3.Crespectively, which indicated that target compound 1 could stabilize h-teloc-myc and c-kit2 G- quaternary DNA, and had a stable effect on h-tclo G-quad DNA. The result of Gaohao. PCR-Stop further proved that target compound 1 could inhibit the PCR amplification products of three kinds of G- quadruplex DNAs at 104m concentration. UV-melting experiment confirmed that target compound 1 could also stabilize the structure of human telomere i-motif DNA, and the target compound 1 could stabilize the structure of human telomere i-motif DNA. But ct-DNA. 2. The results of competitive FRET-melting experiments showed that the% values of Fc-mycT and Fc-kit2T did not change significantly in the presence of 25 times excess double helix ds26, indicating that target compound 1 preferentially binds to the quadruplex. In the presence of 50 times of double helix ds26, the TM value of Fc-kit2T decreased greatly, which indicated that the selectivity of target compound 1 to Fc-kit2T decreased when the concentration of double helix DNA increased. In the presence of 10 times excess double helix ds26, the ATm value of FA-teloT decreased from 9.6 鈩，

本文编号：1885548