【作者】 杨源源；

【导师】 张志超；

【作者基本信息】 大连理工大学 ， 生物化工， 2007， 硕士

【摘要】 研究化合物与生物大分子DNA的相互作用有助于发展新的DNA结构探针、切断剂和抗癌、抗病毒药物，尤其是化合物与DNA的三维空间结构的研究不仅有助于从分子水平上了解抗癌药物的作用机理，而且为设计临床上更为有效的抗癌药物提供理论指导。本论文对本实验室自主合成的一系列8-氧-8H-苊并[1，2-b]吡咯衍生物进行了研究，利用SYBR Green-DNA熔解曲线、紫外-可见光谱、荧光光谱、圆二色光谱等技术手段分析了苊并类化合物与DNA的作用，尤其是利用圆二色光谱技术研究了其与DNA结合的空间几何构型，并且在细胞水平评价了它们的抗肿瘤活性以及它们与DNA的结合构型对其发挥抗肿瘤活性的影响。本论文工作主要分为以下四部分：（1）筛选有DNA嵌入活性的化合物。利用SYBR Green-DNA熔解曲线和圆二色（CD）光谱技术对四十多种苊并类化合物的DNA嵌入能力进行了分析，通过比较化合物的化学结构发现了功能基团。（2）研究在不同条件下有DNA损伤活性的化合物Ⅲ-1与DNA的结合构型。通过DNA损伤实验发现化合物Ⅲ-1：3-（N，N’-二乙基-乙二胺基）-8-氧-8H-苊并[1，2-b]吡咯-9-腈能够切割单链环型M13 mp18 DNA，并且其损伤能力随着剂量的不同而不同；利用分子光谱技术分析了化合物Ⅲ-1和CT DNA在不同的n（Ⅲ-1）／n（CT DNA）比值下的结合模式变化及离子强度、pH、温度等外部条件对Ⅲ-1-DNA体系结合模式的影响。（3）分析化合物与DNA的结合构型与其抗癌活性的关系。分析了九个Ⅲ-1类似物与DNA的结合能力、结合构型以及抗癌活性。研究表明带有甲基哌嗪支链的化合物Ⅷ-3以分子长轴与碱基对长轴平行的方式嵌入到DNA碱基对中，而带有氨基烷基支链的化合物Ⅲ-2和Ⅲ-3则是以垂直方式嵌入到DNA中的。细胞毒性实验表明九个化合物都具有优秀的抗肿瘤活性(IC₅₀=10^-6～10⁽-7(M^-1)，值得一提的是化合物Ⅷ-3除了具有较低的IC₅₀外，还能够引起细胞的凋亡而后者仅能引起细胞坏死，但是它与DNA的亲和性较化合物Ⅲ-2和Ⅲ-3要弱的多。结果表明与人们一般认为的化合物与DNA的亲和性和其抗癌活性呈正相关的观点不同，苊并类化合物与DNA的结合几何模型对其发挥抗癌活性起到了至关重要的作用。（4）探讨苊并类母体化合物与DNA的相互作用。通过对母体结构与DNA的相互作用及其抗癌活性的研究，进一步证明了化合物的碱性支链的引入对其DNA嵌入能力和发挥抗癌活性的发挥起到了重要作用。更多还原

【Abstract】 There has been substantial interest in understanding the DNA binding properties of artificial molecules in the hope of developing novel probes of DNA stucture、DNA-cleaving agent and new therapeutic drugs. Especially, the study on the binding topology between the compound and DNA not only be helpful to understand some diseases and the mechanism by which some drugs acts but also play an important role in obtaining novel antitumor lead compounds.From combination of SYBR Green-DNA melt curve, absorption titration, fluoresence titration and circular dichroism studies, the binding of a series of acenaphtho[1,2-b] pyrrole derivatives synthesized by our lab with CT DNA are analysized. Moreover, binding geometry and consequently the antitumor potency, the possible relationship between the binding mode of ligand-DNA and its antitumor property, and pharmaceutical purpose are reaseached. This thesis consists of four sections as follows:（1） Screen the DNA intercalators. Screen the DNA intercalators from the acenaphthene derivatives by means of SYBR Green-DNA melt curve and circular dichroism spectra. Through comparing the structure of the compounds, the funtional groups are analysized.（2） Elucidate the binding geometry of the DNA cleaverⅢ-1 with DNA. The cleavege abilities are evaluated for the DNA intercalators, the result is found that compoundⅢ-1 is a dose-dependent cleaver, which can cleave the circluar M13 mpl8 DNA into linear DNA. Then the binding geometry ofⅢ-1 to CT DNA are researched by molecular spectroscopy, there are two different interaction mechanisms involved in the whole interaction process depending on the n （Ⅲ-1）/n （CT DNA）. The effect of the ionic strength, pH and temperature on the binding mode is studied.（3） Analyze the relationship between the DNA intercalation geometry and the antitumor properties. The binding affinities, intercalation geometries with DNA and antitumor activities for nineⅢ-1 analogues are investigated in detail. The compounds containing a methylpiperazine substitution （Ⅷ-3） intercalate in a fashion that the long axis of the molecule parallel to the base-pair long axis, while the alkylamine-substituted compounds （Ⅲ-2,Ⅲ-3） locate vertically to the long axis of DNA base pairs. Further, cell-based studies show all the compounds exhibit outstanding antitumor activities against two human tumor cell lines with IC₅₀ ranging from 10^-7 to 10^-6 M^-1. Interestingly, compoundⅧ-3 which binding affinity is one of the lowest but alter DNA conformation most significantly, show much lower IC₅₀ value than other compounds. Moreover, it could induce tumor cells apoptosis, while the compoundsⅢ-2 andⅢ-3 could only necrotize tumor cells. It can be concluded that the geometry of the acenaphtho[1,2-b] pyrrole derivatives with DNA contribute much more to the antitumor property than binding affinity.（4） Study the interaction of the chromophore with DNA. The important role of the basic substitutions is further illustrated through combining the result of binding mode studies and antitumor studies.更多还原