Department of Chemistry, Duke University, 124 Science Drive, Durham, NC 27708, USA.
Department of Biochemistry, Duke University School of Medicine, Durham, NC 27710, USA.
Nucleic Acids Res. 2020 Aug 20;48(14):7653-7664. doi: 10.1093/nar/gkaa585.
Small molecule-based modulation of a triple helix in the long non-coding RNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) has been proposed as an attractive avenue for cancer treatment and a model system for understanding small molecule:RNA recognition. To elucidate fundamental recognition principles and structure-function relationships, we designed and synthesized nine novel analogs of a diphenylfuran-based small molecule DPFp8, a previously identified lead binder of MALAT1. We investigated the role of recognition modalities in binding and in silico studies along with the relationship between affinity, stability and in vitro enzymatic degradation of the triple helix. Specifically, molecular docking studies identified patterns driving affinity and selectivity, including limited ligand flexibility, as observed by ligand preorganization and 3D shape complementarity for the binding pocket. The use of differential scanning fluorimetry allowed rapid evaluation of ligand-induced thermal stabilization of the triple helix, which correlated with decreased in vitro degradation of this structure by the RNase R exonuclease. The magnitude of stabilization was related to binding mode and selectivity between the triple helix and its precursor stem loop structure. Together, this work demonstrates the value of scaffold-based libraries in revealing recognition principles and of raising broadly applicable strategies, including functional assays, for small molecule-RNA targeting.
小分子对长链非编码 RNA 转移相关肺腺癌转录物 1（MALAT1）三螺旋的调节作用已被提出作为癌症治疗的一种有吸引力的途径，也是小分子与 RNA 识别的模型系统。为了阐明基本的识别原则和结构功能关系，我们设计并合成了九种新型二苯并呋喃基小分子 DPFp8 的类似物，DPFp8 是先前鉴定的 MALAT1 的先导结合物。我们研究了识别模式在结合中的作用，并进行了计算机模拟研究，以及亲和力、稳定性和体外酶降解三螺旋之间的关系。具体而言，分子对接研究确定了驱动亲和力和选择性的模式，包括有限的配体灵活性，如通过配体预组织和结合口袋的 3D 形状互补观察到的。差示扫描荧光法的使用允许快速评估配体诱导三螺旋的热稳定性，这与 RNase R 核酸外切酶对该结构的体外降解减少相关。稳定化的幅度与三螺旋与其前体茎环结构之间的结合模式和选择性有关。总之，这项工作证明了基于支架的文库在揭示识别原则方面的价值，并提出了广泛适用的策略，包括针对小分子-RNA 的功能测定。
