Targeting Nucleic Acids with an Integrated Virtual and Actual Screen

我们寻求第二次更新我们的计划，以完善和应用综合虚拟和实际筛选 为发现新的抑制端粒酶的先导化合物提供了平台。本期供资期间的进展 这一时期的工作非常出色，我们已经实现了所提出的大部分具体目标。在下一次融资中 在此期间，我们将专注于发现先导化合物，这些化合物选择性地结合生物学上重要的高等生物。 有序端粒DNA四链体并靶向单链端粒：POT 1界面。端粒酶 被确定为目标超过20年，但没有批准的药物抑制其活性，所以新的 需要采取一些办法。我们将抑制POT 1（保护端粒1），一种端粒酶必需的蛋白质 活性，通过靶向端粒DNA底物，并通过直接靶向POT 1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 筛选平台 核酸仍然是小分子治疗剂的代表性不足的靶标。越来越多的 有证据表明，非B DNA结构在基因表达中起着重要作用，特别是在 端粒的功能针对这些结构要素是一个有吸引力的创新战略， 开发新的治疗药物。由单个DNA形成的高阶DNA四链体结构 人类端粒的一条链仍然是一个非常差的和不适当的研究目标， 生物学方法是不成功的。我们使用了计算建模和 实验生物物理学推导一致的结构模型端粒DNA序列高达192 碱基或八个四重重复。高阶四重堆积 因此，界面代表了端粒DNA小分子稳定化的新靶点。这种稳定化 应该通过将底物DNA锁定在不可用的形式来抑制端粒酶。一个代表性更低的 靶比非B DNA是核酸-蛋白质相互作用，特别是单链DNA：蛋白质 与POT 1一样存在复合物。我们将靶向POT 1的DNA底物和DNA结合功能， 这可能导致端粒酶的协同抑制。这是我们现有资金的自然增长 在此期间，我们针对生物学相关的四链体结构，端粒样高阶四链体 核酸结构和端粒ssDNA结合蛋白POT 1，其对于端粒DNA是必需的 复制的具体的目的是：1）利用整合的端粒四链体靶向更高阶的端粒四链体结构。 虚拟和实际屏幕; 2）表征POT 1与高阶四链体的特定相互作用; 3)使用集成的虚拟和实际屏幕靶向POT 1; 4）表征 高阶G-四链体和POT 1结合剂。 !

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. !