Establishing a molecular system for drug targeting of transcriptional control

描述(由申请人提供)：我们已经确定了一类新的分子受体，可以通过药物靶向来调节基因表达。基因表达调控在分子治疗中有着广泛的应用，包括癌症、心脏病、炎症、神经退行性变、内分泌学、哮喘和糖尿病。受这一发现影响的科学界非常广泛，既有基础科学家，也有翻译科学家。这些分子靶点包括二级DNA结构(G-四链体和I-基序)，广泛存在于c-Myc、Bcl-2、VEGF、Hif-1a、PDGF-A和RET等基因的启动子元件中。存在用小分子稳定这些二级DNA结构导致抑制细胞中基因表达的原理证明，这种抑制依赖于启动子区域完整的G-四联体。夸氟辛是一种源自PI实验室的药物，是一种一流的G-四链相互作用药物，针对核糖体DNA中的G-四链，目前处于第二阶段临床试验。临床试验中的选择性和相应的没有任何严重毒性在一定程度上取决于对癌细胞核仁的选择性摄取，以及四氟新对rDNA中发现的平行型G-四链的400倍选择性。这项提议的目的是提供对消音器元件的结构性洞察，该消音器元件由与消音器元件相对应的双链区域内的G-四链体和I-基序组成。这就是挑战所在，因为I-基序通常只在质粒超螺旋状态下或作为转录径流的结果而产生。我们已经提出了这个问题的解决方案，以及提供基于结构的方法的技术，并开发了一种高通量方法来筛选与这种复合结构相互作用的化合物。在双链分子中形成复合I-基序/G-四链结构的能力将使我们能够识别具有增强靶标选择性的小分子。这项提案所涉及的挑战是设计一个系统，在这个系统中，我们可以从外部控制细胞中基因的生产。这项技术如果成功，将在癌症、心脏病和阿尔茨海默病的治疗中得到应用。

DESCRIPTION (provided by applicant): We have identified a new class of molecular receptors amenable to drug targeting to modulate gene expression. Modulation of gene expression has broad applications in molecular therapeutics, including cancer, heart disease, inflammation, neurodegeneration, endocrinology, asthma, and diabetes. The scientific community affected by this discovery is very broad and includes basic as well as translational scientists. These molecular targets comprise secondary DNA structures (G- quadruplexes and i-motifs) found frequently in the promoter elements of a wide variety of genes including c-Myc, Bcl-2, VEGF, Hif-1a, PDGF-A, and RET. Proof of principle exists that stabilization of these secondary DNA structures with small molecules results in repression of gene expression in cells and this repression is dependent on an intact G-quadruplex in the promoter region. Quarfloxin, a drug originating from the PI's laboratory, is a first-in-class G-quadruplex-interactive drug that targets G-quadruplexes in ribosomal DNA and is in phase II clinical trials. The selectivity and corresponding lack of any serious toxicity in the clinical trials is partially dependent on the selective uptake into the nucleolus of cancer cells along with the 400-fold selectivity of Quarfloxin for the parallel-type G-quadruplex found in rDNA over duplex DNA. The objective of this proposal is to provide structural insight into silencer elements consisting of a G- quadruplex and i-motif within a duplex region that corresponds to the silencer elements. Herein lies the challenge because the i-motif is only normally generated under the negative superhelicity found in a plasmid supercoiled state or as a consequence of transcriptional runoff. We have proposed a solution to this problem along with techniques to provide a structure-based approach and development of a high-throughput assay for screening for compounds that interact with this composite structure. The ability to form the composite i-motif/G-quadruplex structure within a duplex molecule will permit us to identify small molecules that have enhanced target selectivity. The challenge addressed in this proposal is to design a system in which we can externally control the production of genes in cells. This technology, if successful, will have applications in the treatment of cancer, heart disease, and Alzheimer's disease.