DNA ALKYLATION BY THE ANTITUMOR ANTIBIOTIC LEINAMYCIN

莱那霉素是一种破坏dna的天然产物，具有强大的抗癌活性。这种抗生素结构独特，代表了一类新的dna损伤抗生素。由于leinamycin具有强大的抗癌活性，并通过其新的化学机制破坏DNA，因此对这种抗生素的研究具有实用和基础的意义。leinamycin的DNA损伤是由抗生素与硫醇反应引起的，最近的研究表明该化合物可引起氧化性和烷基化性DNA损伤。DNA-烷基化化学是本研究的重点，它通过巯基介导的leinamycin重排产生亲电的episulium离子，使鸟嘌呤残基N7处的双链DNA烷基化。虽然我们已经初步掌握了leinamycin破坏DNA的机制，但我们对这种抗生素与DNA相互作用的化学反应和分子识别过程的理解还远远不够。本提案中描述的工作调查了leinamycin-DNA相互作用的几个新方面，并将对这种不寻常的抗生素如何有效地烷基化DNA提供更深入的了解。提出的工作的具体目的如下：(1)表征Leinamycin-DNA加合物。其中包括研究leinamycin对DNA烷基化的序列特异性，分析leinamycin-guanosine加合物的稳定性，以及寻找新的leinamycin-DNA加合物（除了N7-guanosine）。(2)研究Leinamycin的“替代”（非硫醇激活）激活模式。迄今为止，莱那霉素被认为是一种硫醇依赖的DNA损伤剂。我们计划研究几种替代的，非硫醇激活模式的DNA烷基化激活的leinamycin，我们已经发现在我们最近的工作过程中。(3) Leinamycin检测非共价DNA关联。我们已经获得了初步的证据，leinamycin与DNA非共价结合。抗生素的结构特征是重要的DNA结合，DNA结合模式和雷那霉素对双链DNA的亲和力将被检查。

Leinamycin is a DNA-damaging natural product with potent anticancer activity. This antibiotic is structurally unique and represents a new class of DNA-damaging antibiotics. Because of its potent anticancer activity and the novel chemical mechanisms by which leinamycin damages DNA, studies of this antibiotic are of both practical and fundamental interest. DNA damage by leinamycin is triggered by reaction of the antibiotic with thiols and recent work has shown that the compound causes both oxidative and alkylative DNA damage. The DNA-alkylation chemistry that is the focus of this proposal occurs via a surprising thiol-mediated rearrangement of leinamycin that yields an electrophilic episulfonium ion which alkylates double-stranded DNA at N7 of guanine residues. Although we have a preliminary grasp on the mechanisms by which leinamycin damages DNA, our understanding of the chemical reactions and molecular-recognition processes involved in the interaction of this antibiotic with DNA is far from complete. The work described in this proposal investigates several novel aspects of leinamycin-DNA interactions and will provide a deeper understanding of how this unusual antibiotic efficiently alkylates DNA. The Specific Aims of the proposed work are as follows: (1) Characterize Leinamycin-DNA adducts. The includes investigation of the sequence-specificity of DNA alkylation by leinamycin, analysis of the stability of leinamycin-guanosine adducts, and searches for new leinamycin-DNA adducts (other than N7-guanosine). (2) Investigate "Alternate" (Non Thiol-Activated) Modes of Leinamycin Activation. Leinamycin has thus far been characterized as a thiol-dependent DNA damaging agent. We plan to investigate several alternate, non thiol-activated modes for the activation of DNA alkylation by leinamycin that we have discovered during the course of our recent work. (3) Examine Non-Covalent DNA Association by Leinamycin. We have obtained preliminary evidence that leinamycin non-covalently associates with DNA. The structural features of the antibiotic that are important for DNA binding, the mode of DNA association and the affinity of leinamycin for duplex DNA will be examined.