Design and study of artificial nucleases for DNA photocleavage

DNA光裂解人工核酸酶的设计与研究

• 批准号: 341612-2007
• 负责人: McFarland, Sherri
• 金额: $ 1.81万
• 依托单位: Acadia University
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2007
• 资助国家: 加拿大
• 起止时间: 2007-01-01 至 2008-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=361368
• 关键词: Design; study; artificial; nucleases; DNA

Artificial nucleases are molecules capable of inducing single- or double-strand breaks in the DNA backbone.  Molecules that cleave DNA  upon activation with light are especially important in photodynamic therapy (PDT), a strategy that has been applied to the successful treatment of certain cancers and age-related macular degeneration.  Current clinical agents for PDT suffer from three major limitations:  (i) requirement for oxygen to function, (ii) minimum absorption of light at wavelengths most transparent to tissue, and (iii) inability to produce double-strand breaks.  This proposal outlines a research plan that is devoted to overcoming such limitations by introducing new  agents that elicit double-strand breaks in DNA under hypoxic conditions when activated with 700-900 nm light.  The first proposed family of DNA photocleavers is based on dinuclear Ru(II) complexes that absorb light >700 nm.  Extension of this family will include incorporation of Rh(II) to make hybrid multimetallic systems that cleave DNA in the absence of oxygen.  The multimetallic framework will provide two sites of reactivity for DNA cleavage.  Dual reactivity within 15-16 base pairs will introduce double-strand breaks which are more difficult for the cellular machinery to repair.       A related research stream involves  the jadomycins, a family of secondary metabolites that induces DNA strand scissions in the presence of Cu(II) or with light activation.  While  they fall short of the properties outlined for the ideal PDT agent, jadomycins exhibit cytotoxic activity against both Gram positive and Gram negative bacteria as well as some drug-resistant cancer cell lines. This research program will determine whether direct DNA cleavage contributes to the overall cytotoxicity of the jadomycins at intracellular Cu(II) concentrations.  In addition, the DNA photocleaving properties of the jadomycins will be investigated, with particular attention on the mechanism that predominates with Cu(II) activation versus light activation.  The role that L-digitoxose, a structural feature of jadomycin B, plays in DNA cleavage will also be probed to better understand how glycone incorporation aids in DNA recognition.

人工核酸酶是能够在DNA主干中诱导单链或双链断裂的分子。在光激活下切割DNA的分子在光动力疗法（PDT）中尤为重要，这一策略已被应用于成功治疗某些癌症和年龄相关性黄斑变性。目前用于PDT的临床药物存在三个主要限制：(i)需要氧气才能发挥作用；（ii）对组织最透明波长的光吸收最小；（iii）不能产生双链断裂。该提案概述了一项研究计划，该计划致力于通过引入新的药物来克服这些限制，这些药物在缺氧条件下被700-900 nm的光激活时引发DNA双链断裂。第一个被提出的DNA光切割器家族是基于双核Ru（II）配合物的，它吸收波长为700 nm的光。这个家族的延伸将包括Rh（II）的结合，以制造在缺氧条件下切割DNA的杂交多金属系统。多金属骨架将为DNA切割提供两个反应活性位点。15-16个碱基对内的双反应性会导致双链断裂，这对细胞机器来说更难以修复。一个相关的研究流涉及到jadomycin，这是一类次级代谢产物，在Cu（II）存在或光激活下诱导DNA链断裂。虽然它们没有理想的PDT试剂的特性，但jadomycin对革兰氏阳性和革兰氏阴性细菌以及一些耐药癌细胞系都表现出细胞毒性活性。该研究计划将确定在细胞内Cu（II）浓度下，直接的DNA切割是否有助于贾霉素的整体细胞毒性。此外，还将研究jadomycin的DNA光切割特性，特别关注Cu（II）激活与光激活的主要机制。L-digitoxose是jadomycin B的一个结构特征，它在DNA切割中的作用也将被探讨，以更好地了解糖苷结合如何辅助DNA识别。