Programmable peptide-guided protein degradation

可编程肽引导的蛋白质降解

• 批准号: 10741655
• 负责人: Pranam Chatterjee
• 金额: $ 38.77万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10741655
• 关键词: 2019-nCoV; Active Sites; Address; Algorithms; Amino Acid Sequence; Antibodies; Architecture; Base Sequence; Binding; Biological Assay; CRISPR/Cas technology; Cells; Chemistry; Chimera organism; Chimeric Proteins; Code; Complex; Consumption; DNA; Data Set; Databases; Development; Disease; Employment; Encapsulated; Engineering; Fibronectins; Fluorescence; Formulation; Foundations; Gene Delivery; Generations; Genetic; Goals; Human; In Vitro; Laboratories; Language; Learning; Length; Libraries; Malignant Neoplasms; Masks; Mediating; Messenger RNA; Methodology; Methods; Modality; Modeling; Molecular Conformation; Mutagenesis; Output; Pathogenicity; Peptides; Phosphoric Monoester Hydrolases; Phosphotransferases; Positioning Attribute; Post-Translational Protein Processing; Protac; Protein Engineering; Protein Isoforms; Proteins; Proteome; Protocols documentation; RNA; Rapid screening; Research; Specific qualifier value; Structure; Techniques; Technology; Testing; Therapeutic; Training; Translating; Translations; Ubiquitin-Proteasomal Pathway; Validation; Variant; Work; design; disease phenotype; experimental analysis; gene therapy; grasp; in silico; in vivo; in vivo evaluation; interest; iterative design; lipid nanoparticle; mutant; nanobodies; novel therapeutic intervention; novel therapeutics; pharmacologic; protein aminoacid sequence; protein complex; protein degradation; protein protein interaction; rapid testing; scaffold; screening; small molecule; small molecule inhibitor; tool; ubiquitin-protein ligase

Summary Over 600 human proteins have been recently prioritized as key cancer targets, with nearly half being considered ‘intractable’ by standard small-molecule inhibition approaches, due to target instability and active site accessibility constraints. By redirecting the ubiquitin-proteasomal pathway (UPS) for targeted protein degradation, the proteolysis-targeting chimera (PROTAC) technology provides a potential solution, enabling rapid and continuous target consumption as well as the stronger pharmacological effects than small molecule inhibition. Nonetheless, PROTACs suffer from similar developmental hurdles as small molecules and cannot be easily designed for motif or post-translational modification-specific targeting. To address these hurdles, research efforts have shifted toward gene therapy approaches by introducing the concept of protein-mediated protein degradation. Here, E3 ubiquitin ligases are redirected by replacing their natural substrate binding domains with “off-the-shelf” binding domains, including nanobodies, antibodies, and DARPins, to generate target-specific ubiquibodies. To augment this platform, we recently exploited natural protein-protein interaction information to develop algorithmic pipelines that prioritize target-selective peptides which can be fused to the E3 ubiquitin ligase conjugation domains to induce target protein degradation. In this project, we will augment our current methods to enable the development of these ubiquibodies (uAbs) for any protein, including those deemed ‘intractable’ by small molecule-based means. To do this, we will automate a bipartite algorithmic pipeline that leverages recent advancements in protein language modeling as well as existing co-complex databases to design peptide binders to diverse protein targets, including those with solved co-crystals as well as those with minimal structural information. Specifically, our pipeline will take user-specified target proteins as inputs, and generate prioritized lists of candidate peptide binders as outputs, enabling subsequent generation of uAbs for target degradation. Through library-on-library fluorescence-based assays in human cells and subsequent encapsulation of uAb mRNA in lipid nanoparticles (LNPs), we will develop a scalable method to test and translate our degraders for downstream in vivo validation. In total, this work will generate a robust peptide design tool that will enhance targeted protein degradation efforts and lay the foundation for programmable proteome editing.

概括 最近将超过600种人类蛋白作为关键癌症的优先级优先考虑，几乎一半是 由于目标不稳定性和主动性，标准的小分子抑制方法被认为是“棘手的” 站点可访问性约束。通过重定向靶向蛋白的泛素 - 蛋白酶体途径（UPS） 退化，靶向蛋白水解嵌合体（Protac）技术提供了潜在的解决方案，使得能够 与小分子相比，快速，连续的目标消耗以及更强的药物效应 抑制。但是，Protac遭受与小分子相似的发展障碍，不能是 易于设计用于基序或翻译后修改特定的靶向。要解决这些障碍， 研究工作已通过引入蛋白质介导的概念转向基因治疗方法 蛋白质降解。在这里，E3泛素连接酶通过更换其自然底物结合而被重定向 具有“现成”绑定域的域，包括纳米动物，抗体和DARPINS，生成 目标特异性的泛质。为了增加该平台，我们最近探索了天然蛋白质 - 蛋白质相互作用 信息开发算法管道的信息，该管道优先考虑目标选择性辣椒，可以融合到 E3泛素连接酶结合结构域诱导靶蛋白降解。在这个项目中，我们将增加 我们目前的方法可以为任何蛋白质的泛质开发（UABS）的开发，包括 基于小分子的均值被认为是“棘手的”。为此，我们将自动化两分算法 利用蛋白质语言建模以及现有共同复合的最新进展的管道 数据库以设计肽粘合剂针对各种蛋白质靶标，包括具有解决共晶的蛋白质靶标 作为最小结构信息的人。具体而言，我们的管道将将用户指定的目标蛋白作为 输入，并生成优先级的候选肽粘合剂列表作为输出，从而使后续生成 UAB的目标降解。通过在人类细胞中基于上图书馆的荧光测定法和 随后在脂质纳米颗粒（LNP）中对UAB mRNA的封装，我们将开发一种可扩展的方法 测试并翻译我们的降级器以进行体内验证下游。总的来说，这项工作将产生强大的 肽设计工具将增强靶向蛋白质降解努力并为 可编程蛋白质组编辑。