Novel Approaches for PROTAC Drug Discovery

PROTAC 药物发现的新方法

• 批准号: 10081299
• 负责人: Karteek Kadimisetty
• 金额: $ 39.71万
• 依托单位: LIFESENSORS, INC.
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-15 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10081299
• 关键词: Affinity; Amyloid beta-Protein; Antineoplastic Agents; Autophagocytosis; Autophagosome; Binding; Biological Assay; Cell model; Cells; Cellular Assay; Clinic; Communities; Consumption; Cullin Proteins; Cultured Cells; Development; Disease; Drug Targeting; Drug vehicle; Enzymes; Family; Glues; Goals; Health; Human Genome; Immunity; Immunomodulators; Lead; Ligase; Link; Location; Lysosomes; MDM2 gene; Measures; Mediating; Metabolism; Methods; Modality; Modeling; Molecular; Monitor; Morphologic artifacts; Neurodegenerative Disorders; Neurofibrillary Tangles; Neurons; Pattern; Penetrance; Pharmaceutical Preparations; Pharmacologic Substance; Phase; Play; Polymers; Process; Protac; Proteins; Research; Research Personnel; Revlimid; Role; Signal Transduction; Site; Specificity; Structure-Activity Relationship; System; T-Lymphocyte; Technology; Testing; Thalidomide; Time; Ubiquitin; Ubiquitination; Uncertainty; United States Food and Drug Administration; Western Blotting; Work; analog; base; cancer therapy; chimera drug; design; drug discovery; immune function; innovation; member; misfolded protein; multicatalytic endopeptidase complex; mutant; novel; novel strategies; novel therapeutics; overexpression; preclinical development; protein aggregation; protein degradation; protein transport; side effect; small molecule; tau Proteins; tau aggregation; transcription factor; ubiquitin ligase; ubiquitin-protein ligase

PROteolysis TArgeting Chimeras (PROTACs) is a new therapeutic class comprised of small molecules binding a target protein and a ubiquitin (Ub) E3 ligase, enabling selective target degradation. PROTACs’ advantages include exquisite selectivity, tolerance of weak binders, and maximal degradation with limited target engagement. The first PROTACs employed the E3 ligase component pVHL to degrade target proteins. Around the same time, thalidomide and related analogs (IMiDs) were successfully repurposed as anti-cancer agents, and subsequently their ubiquitin ligase-associated molecular mechanism was discovered, when thalidomide was shown to bind to cereblon (CRBN), a pVHL-like subunit of a Cullin 4-type ubiquitin ligase. IMiDs promote interaction of this E3 ligase transcription factors that control T cell immunity. In cells, Ub-mediated signaling regulates protein content, location, and activity, primarily through protein degradation, and dysregulation of ubiquitin ligases is linked to numerous devastating diseases. Thus, ligases are promising drug targets and vehicles for PROTACs. Of ~700 Ub ligases, only CRBN and pVHL have been exploited for PROTAC development, a process hindered by several issues. There is a disconnect between the rapid synthesis of new PROTACs by chemists and the time- consuming, artifact-susceptible immunoblot cell assays used to evaluate them. Moreover, binding of a PROTAC to its target does not ensure degradation, owing to steric hindrance, ubiquitylation at the wrong site or in the wrong chain configuration (K63 vs K48), or metabolism inside or poor penetrance into cells. Thus, it is difficult for PROTAC chemists to develop meaningful structure activity relationships, which are essential for preclinical development. It is proposed here to develop ligase-selective, high-throughput cellular assays for PROTAC- mediated ubiquitylation of target proteins. In phase I, LifeSensors will employ unique affinity matrices called TUBEs (Tandem Ubiquitin Binding Entities) in a high throughput mode to analyze ubiquitylation patterns of endogenous proteins in cells. This approach offers the potential to expedite discovery of novel PROTACs, establish a relationship between ubiquitylation and degradation, eliminate low throughput, time-consuming western blot analysis, and lead to the timely identification and development of novel PROTAC drugs, as medicinal chemists will be able to design PROTACs efficiently and rationally, eventually encompassing both degradative and non-degradative ubiquitylation. Phase I will be accomplished by establishing a clear relationship between PROTAC-mediated ubiquitylation and degradation for CRBN and HDM2 ligase target proteins in cells and adapting LifeSensors’s TUBEs technology to monitor PROTAC ubiquitylation and degradation in a high throughput fashion in cells. In Phase II the technology will be expanded to entire ligase families.

针对嵌合体的蛋白分解(PROTACs)是一种由小分子结合组成的新的治疗类别 一个目标蛋白和一个泛素(Ub)E3连接酶，能够选择性地降解目标。PROTAC的优势 包括精致的选择性，对弱结合剂的耐受性，以及在有限的靶结合下最大限度地降解。 第一批PROTACs利用E3连接酶组分pVHL降解目标蛋白。差不多在同一时间， 沙利度胺及其类似物(IMids)被成功地重新用作抗癌药物，随后 当沙利度胺被证明与其结合时，其泛素连接酶相关的分子机制被发现 Cereblon(CRBN)，库林4型泛素连接酶的pVHL样亚单位。IMID促进这一E3的互动 控制T细胞免疫的连接酶转录因子。在细胞中，Ub介导的信号调节蛋白质含量， 定位和活性，主要是通过蛋白质降解，泛素连接酶的失调与 无数毁灭性的疾病。因此，连接酶是PROTACs很有前途的药物靶点和载体。约700人 UB连接酶，只有CRBN和pVHL被用于PROTAC的发育，这一过程受到几个 问题。化学家快速合成新的PROTAC与时间之间存在脱节- 消耗的，人工制品敏感的免疫印迹细胞分析用于评估它们。此外，PROTAC的绑定 不能确保降解，由于空间位阻，泛素化在错误的位置或 错误的链构型(K63和K48)，或者在细胞内代谢或进入细胞的能力差。因此，这是困难的 对于PROTAC化学家来说，开发有意义的结构活性关系，这对于临床前是必不可少的 发展。建议建立连接酶选择性、高通量的PROTAC-细胞检测方法。 介导的靶蛋白泛素化。在第一阶段，生命传感器将使用唯一的亲和力矩阵，称为 管(串联泛素结合实体)在高通量模式下分析泛素化模式 细胞内的内源性蛋白质。这种方法提供了加速发现新的PROTAC的可能性， 建立泛素化和降解之间的关系，消除低吞吐量、耗时 Western印迹分析，并导致及时识别和开发新的PROTAC药物，AS 药物化学家将能够有效和合理地设计PROTAC，最终将两者都包括在内 降解性和非降解性泛素化。第一阶段将通过建立明确的关系来完成 PROTAC介导的泛素化与细胞内CRBN和Hdm2连接酶靶蛋白降解的关系 采用Lifesensors的管技术来监测PROTAC的泛素化和高水平的降解 以单元为单位的吞吐量方式。在第二阶段，这项技术将扩展到整个连接酶家族。