Synthesis and DNA binding affinity evaluation of novel gilvocarcin-C-glucosides

描述（由申请人提供）：许多已知与双链DNA结合的天然产物具有两个关键的结构特征，允许与细胞靶标紧密结合：插入DNA主链的平面芳香苷元，以及与主要和/或次要凹槽参与非共价相互作用的一个或多个碳水化合物部分。c -芳基糖苷，在芳香插入物和一个或多个碳水化合物残基之间含有碳-碳键的物质，因此可能被视为开发高亲和力dna结合剂的有希望的候选者。虽然有研究表明，含有附加糖基残基的芳香插入物与缺乏此类残基的插入物相比，具有更强的结合DNA的能力，但文献中还没有系统的研究定量地比较含有一个、两个或三个附加糖基单元的插入物的DNA结合亲和力。在本研究中，我们将制备一系列gilvocarcin M发色团的单、双和三- c -芳基葡萄糖苷衍生物，并通过荧光和紫外光谱技术评估它们与双链DNA的结合常数。c -芳基糖苷天然产物的gilvocarcin家族成员是进行此类研究的理想模板，因为它们是已知的具有高抗肿瘤活性和低总毒性的dna结合剂。该合成计划要求将受保护的碳水化合物、萘和邻碘苯甲酸亚基聚合在一起，并允许在七到九个步骤中制备四种gilvocarcin c -葡萄糖苷衍生物。芳基金属（芳基锂或芳基铜酸盐）试剂与糖硫代内酯的直接偶联是芳烃和碳水化合物之间关键的碳-碳键形成反应，然后是中间半缩酮的立体选择性自由基还原。我们提出的结合研究测量了我们的gilvocarcin衍生物与小牛胸腺DNA、poly(dA/dT)7poly（dA/dT）和poly(dG/dC)7poly（dG/dC）的相互作用强度，这不仅使我们能够评估额外的碳水化合物附着在gilvocarcin发色团上改变DNA结合亲和力的程度，还将使我们能够建立碳水化合物附着在发色团上的位置（对应于在主要和/或次要凹槽中的位置）与DNA结合亲和力之间的相关性；此外，我们的衍生品的DNA序列偏好也将从所获得的数据中显而易见。由于假设c -芳基糖苷天然产物的DNA序列特异性取决于糖苷取代基的性质，因此这些研究将为最终制备含有各种不同碳水化合物的gilvocarcin衍生物铺平道路，这些衍生物有助于建立序列依赖性DNA结合配体的设计范例。这些分子，如果能够选择性地与目标基因的启动子或编码区相互作用，可能最终用于基因转录的调节和作为广泛疾病状态的治疗剂。

DESCRIPTION (provided by applicant): Numerous natural products that are known to bind duplex DNA possess two key structural characteristics that allow tight binding interactions with their cellular target: a planar aromatic aglycone that intercalates the backbone of DNA, and one or more carbohydrate moieties that participate in non-covalent interactions with the major and/or minor grooves. C-Aryl glycosides, substances containing carbon-carbon bonds between an aromatic intercalator and one or more carbohydrate residues, may thus be viewed as promising candidates for the development of high affinity DNA-binding agents. Although it has been shown that aromatic intercalators containing attached glycosyl residues have a superior ability to bind DNA in comparison to intercalators lacking such residues, there exist in the literature no systematic studies quantitatively comparing the DNA binding affinity of intercalators containing one, two, or three appended glycosyl units. In the proposed research, we will prepare a series of mono-, bis-, and tris-C-aryl glucoside derivatives of the gilvocarcin M chromophore and evaluate their association constants for duplex DNA by fluoresence and ultraviolet spectrocopic techniques. Members of the gilvocarcin family of C-aryl glycoside natural products are ideal templates for undertaking such a study since they are known DNA-binding agents that possess high antitumor activity with low overall toxicity. The synthesis plan calls for the convergent assembly of protected carbohydrate, naphthalene, and o-iodobenzoic acid subunits, and will allow the preparation of four gilvocarcin C-glucoside derivatives in seven to nine steps. The key carbon-carbon bond-forming reactions between aromatic and carbohydrate moieties will involve the direct coupling of arylmetal (aryllithium or arylcuprate) reagents and sugar thiolactones, followed by stereoselective radical reduction of the intermediate hemithioketal. The proposed binding studies measuring the strength of interaction of our gilvocarcin derivatives with calf-thymus DNA, poly(dA/dT)7poly(dA/dT), and poly(dG/dC)7poly(dG/dC) will not only allow us to assess the degree to which the attachment of additional carbohydrates to the gilvocarcin chromophore alters DNA binding affinity, but also will allow us to establish a correlation between the site of carbohydrate attachment on the chromophore (corresponding to placement in the major and/or minor grooves) and DNA binding affinity; furthermore, the DNA sequence preferences of our derivatives will also be apparent from the data obtained. Since it has been hypothesized that the DNA sequence specificity of C-aryl glycoside natural products depends on the nature of the glycosidic substituents, these studies will pave the way for the eventual preparation of gilvocarcin derivatives bearing a variety of different carbohydrate moieties useful in establishing paradigms for the design of sequence-dependent DNA-binding ligands. Such molecules, if capable of selectively interacting with the promoters or coding regions of target genes, may ultimately be used for the regulation of gene transcription and as therapeutic agents for a wide range of disease states. PUBLIC HEALTH RELEVANCE: The development of high-affinity DNA-binding molecules is a crucially important goal in the context of current approaches to cancer therapy; indeed, to maximize tumor cell-specific cytotoxicity, drug molecules must be able to bind tightly to unique targets identified within cancer cells. The proposed research will involve the synthesis and DNA binding-affinity evaluation of a series of mono-, bis-, and tris- C-aryl glucoside derivatives of the gilvocarcins, a class of non-toxic antitumor antibiotics viewed as templates for the design of gene-specific cytotoxic agents. Through this study we hope to establish a correlation between the strength of small molecule-DNA interactions and both the number and position of attachment of carbohydrate residues around an aromatic intercalator.