G-Quadruplexes as Targets for Drug Design

某些DNA序列具有形成DNA二级结构的能力，这些二级结构具有与双链DNA截然不同的分子识别特征。我们特别感兴趣的是一组四链结构，叫做g -四联体或DNA四联体。形成g -四联体的基序在真核生物基因组中似乎很少见，但在具有生物学意义的区域（如端粒和启动子区域）中发现。我们已经确定了靶向这些序列的小分子，并影响G-四联体的形成和解绕速率。在本提案中，我描述了将确定这些小分子对G-四联体在癌基因调控区域的开关机制和端粒区域稳定t环中所起作用的影响的实验。g -四重复合物在重组中的其他可能作用及其在体内的存在也将被评估。生物物理方法，包括核磁共振和PAGE，将主要用于本研究。本提案的具体目的是：(1)确定导致g -四重体和i-基序相互作用化合物下调c-myc的分子机制；(2)确定t环侵入复合物的分子结构，并以此为指导开发新的端粒相互作用化合物；(3)确定g -四重体相互作用化合物对其他蛋白质- g -四重体复合物的影响，包括拓扑异构酶i - g -四重体DNA；(4)利用LM-PCR检测g -四重结构在体内的发生情况。

Certain DNA sequences have the capability to form DNA secondary structures that have molecular recognition features quite distinct to duplex DNA. In particular, we are interested in a group of four-stranded structures called G-quadruplexes or DNA tetraplexes. Motifs for the formation of G-quadruplexes appear to be rare in the eukaryotic genome but are found in regions of biological significance (e.g., telomeres and promoter regions). We have identified small molecules that target these sequences and affect the rate of formation and unwinding of G- quadruplexes. In this proposal, I describe experiments that will determine the effect of these small molecules on the role that G- quadruplexes play in switch mechanisms in regulatory regions of oncogenes and in stabilizing t-loops in telomeric regions. Other possible roles of G-quadruplexes in recombination and their in vivo existence will also be evaluated. Biophysical methods, including NMR and PAGE, will be used primarily in this study. The specific aims of this proposal are to (1) determine the molecular mechanism that results in the down- regulation of c-myc by G-quadruplex and i-motif-interactive compounds; (2) determine the molecular structure of the t-loop invasion complex and use this as a guide to develop new telomere- interactive compounds; (3) determine the effect of G-quadruplex-interactive compounds on other protein-G-quadruplex complexes, including the topoisomerase I-G-quadruplex DNA; (4) use LM-PCR to probe for the occurrence of G-quadruplex structures in vivo.