Targeting Nucleic Acids with an Integrated Virtual and Actual Screen

我们为我们的计划寻求第二次更新，以完善和应用虚拟和实际相结合的筛查 发现抑制端粒酶的新先导化合物的平台。在当前供资期间取得的进展 这一时期表现突出，我们已经实现了提出的大部分具体目标。在下一次资助期间 期间，我们将专注于发现选择性地与具有生物重要性的更高- 订购端粒DNA四链并靶向单链端粒：POT1接口。端粒酶有 20多年来一直被确定为靶点，但还没有批准的药物来抑制其活性，所以新的 方法是必需的。我们将抑制POT1(端粒保护1)，这是一种端粒酶必不可少的蛋白质 活性，通过靶向端粒DNA底物和直接靶向POT1端粒DNA结合 功能。我们现在提出的基础研究将继续发展和完善我们的综合 放映平台。 核酸仍然是小分子治疗剂的目标。有越来越多的人 证据表明，非B-DNA结构在基因表达中发挥着重要作用，特别是在 端粒的功能。以这些结构要素为目标是一种有吸引力的创新战略 开发新的治疗剂。由单链形成的高阶DNA四链结构 人类端粒的链仍然是一个非常糟糕和不正确的研究目标，因为传统的结构 生物学方法一直不成功。我们结合使用了计算建模和 实验生物物理学为高达192的端粒DNA序列推导出一致的结构模型 基数，或八个四倍体重复，在当前赠与期。高阶四路叠加 因此，界面代表了端粒DNA小分子稳定的新靶点。这种稳定 应该通过将底物DNA锁定在不能使用的形式来抑制端粒酶。一个代表更少的人 靶点比非B DNA是核酸与蛋白质的相互作用，尤其是单链DNA：蛋白质 复合体，与POT1一样存在。我们将针对POT1的DNA底物和DNA结合功能， 这可能导致对端粒酶的协同抑制。这是我们目前资金的自然发展 我们以生物相关的四链结构为目标的时期，到类似端粒的高阶四链结构 端粒DNA所必需的核酸结构和端粒单链DNA结合蛋白POT1 复制。具体的目标是：1)以更高阶端粒四链结构为靶点 虚拟和实际屏幕；2)表征POT1与高阶四联体的特定相互作用； 3)使用集成的虚拟和实际筛选来靶向POT1；4)表征POT1的生物学特性 高阶G-四链体和POT1结合剂。 好了！

We seek a second renewal for our program to refine and apply an integrated virtual and actual screening platform for the discovery of new lead compounds to inhibit telomerase. Progress during the current funding period was outstanding, and we have achieved most of the specific aims proposed. During the next funding period, we will focus on the discovery of lead compounds that bind selectively to biologically important higher- order telomeric DNA quadruplexes and targeting the single strand telomere:POT1 interface. Telomerase has been identified as a target for over 20 years, but there are no approved drugs that inhibit its activity, so new approaches are required. We will inhibit POT1 (Protection of Telomere 1), a protein essential for telomerase activity, by targeting the telomeric DNA substrate, and by directly targeting the POT1 telomeric DNA binding function. We now propose fundamental studies that will continue to develop and refine our integrated screening platform. Nucleic acids remain underrepresented targets for small molecule therapeutic agents. There is mounting evidence to indicate that non-B DNA structures play prominent roles in gene expression and especially in the function of telomeres. Targeting these structural elements is an attractive and innovative strategy for the development of new therapeutic agents. The higher-order DNA quadruplex structures formed by the single strand of the human telomere remain a very poorly and improperly investigated target as traditional structural biology approaches have been unsuccessful. We have used a combination of computational modeling and experimental biophysics to derive consistent structural models for telomeric DNA sequences of up to 192 bases, or eight quadruplex repeats, in the current grant period. The higher-order quadruplex stacking interfaces thus represent a new target for small molecule stabilization of telomeric DNA. This stabilization should inhibit telomerase by locking the substrate DNA in an unusable form. An even more under represented target than non-B DNA is the nucleic acid-protein interaction, particularly single stranded DNA:protein complexes, as exist with POT1. We will target both the DNA substrate and the DNA-binding function of POT1, which could lead to synergistic inhibition of telomerase. This is a natural progression from our current funding period where we targeted biologically relevant quadruplex structures, to telomere-like higher-order quadruplex nucleic acid structures and the telomere ssDNA binding protein POT1, which is essential for telomeric DNA replication. The Specific Aims are: 1) Targeting higher-order telomeric quadruplex structures using integrated virtual and actual screens; 2) Characterizing the specific interactions of POT1 with higher-order quadruplexes; 3) Targeting POT1 using integrated virtual and actual screens; 4) Characterizing the biological properties of higher-order G-quadruplexes and POT1 binding agents. !