Targeting the IKK-Binding Domain of NEMO for Inhibitors Discovery

摘要 核因子-κB必需调节剂(NEMO)是激活细胞外信号转导通路的重要组成部分 典型的NF-κB途径及其通过募集κB激酶抑制因子IKKα发挥作用 和IKKβ到IKK综合体。抑制IKKα和IKKβ激酶的相互作用 而NEMO被认为是抑制NF-κB的一种有吸引力的治疗途径，因为 通路在人类疾病中的作用，包括炎症性和自身免疫性疾病以及 癌症。该策略被证明是有效的利用多肽，对应于 IKKS的Nemo结合域(NBD)作为IKK复合体抑制物的体内外研究 疾病的模型。该项目的总体目标是了解 NEMO抑制及其在小分子抑制剂开发中的应用。长的- 项目的期望值是为缓蚀剂的设计和开发奠定结构基础。 要求高亲和力结合和特异性。该项目将经过三个阶段的发展 具体目标：1.将利用结构和计算方法来表征 依赖于IKK结合新结构的NEMO-抑制剂络合物的结构 NEMO的区域，很容易产生X射线结构。此外，可以使用的结构 封闭和半开放结构中的未结合NEMO将被用于缓蚀剂设计。2. NEMO结构将指导多肽和小分子抑制剂的合理设计，以 从NBD发展成为具有独特物理和化学性质的有效缓蚀剂 检测不同结合部位的潜在抑制作用。3.尼莫之间的相互作用 和抑制剂将通过生化、生物物理和细胞方法进行表征，以 量化结合亲和力、验证结合、识别结合部位并表征 装订方式和要求。此外，细胞模型系统中的生物学评价 依赖于NF-κB信号生存的癌症，将为 未来的临床前发展和临床研究。相关调查人员的专业知识 在研究中符合该项目的多学科特征，并具有分析经验 发展和生物物理表征、基于核磁共振的筛选和结构 测定、X射线结晶学、多肽化学和合成药物化学， 癌症生物学和抗癌药物，以及在NEMO表征方面的长期经验。 这项研究的结果将进一步加深我们对需求和 NEMO抑制的后果并为开发新的治疗方法开辟了道路 人类疾病。

Abstract The NF-κB essential modulator (NEMO) is an essential component in the activation of the canonical NF-κB pathway and exerts its function by recruiting the inhibitor of κB kinases IKKα and IKKβ to the IKK complex. Inhibition of the interaction between the IKKα and IKKβ kinases and NEMO is considered an attractive therapeutic avenue to the inhibition of NF-κB, due to the pathway’s role in human diseases, encompassing inflammatory and autoimmune diseases and cancer. The strategy was shown to be effective utilizing a peptide, corresponding to the NEMO-binding domain of the IKKs (NBD), as an IKK complex inhibitor in in vitro and in vivo models of disease. The overall goal of the project is the understanding of the mechanism for NEMO inhibition and its exploitation for the development of small molecule inhibitors. The long- term expectation of the project is to establish the structural basis for inhibitor design and the requirements for high affinity binding and specificity. The project will evolve through three specific aims: 1. Structural and computational methods will be utilized to characterize the structure of NEMO-inhibitor complexes, which rely on a new construct of the IKK-binding domain of NEMO which easily yields X-ray structures. Furthermore, the available structures of unbound NEMO in a closed and semi-open structure will be utilized for inhibitor design. 2. The NEMO structures will guide the rational design of peptide and small molecule inhibitors, to develop into potent inhibitors with distinct physical and chemical properties from the NBD and testing the potential of different binding sites for inhibition. 3. The interaction between NEMO and inhibitors will be characterized by biochemical, biophysical and cellular methods, to quantify the binding affinity, validate binding, identify the binding site and characterize the binding mode and requirements. In addition, the biological evaluation in cellular model systems of cancer that rely on NF-κB signaling for survival, will provide the rationale and justification for future preclinical development and clinical studies. The expertise of the Investigators involved in the research matches the multidisciplinary character of the project, with experience in assay development and biophysical characterization, NMR-based screening and structure determination, X-ray crystallography, peptide chemistry and synthetic medicinal chemistry, cancer biology and anticancer drugs, and a long experience in the characterization of NEMO. The outcome of this research will further our understanding of the requirements and consequences of NEMO inhibition and open the way to the development of new therapies for human disease.