Targeted Gene Regulation Using Engineered Synthetic Transcriptional Regulators

使用工程合成转录调节剂进行靶向基因调节

• 批准号: 10538129
• 负责人: Colin Stuart Swenson
• 金额: $ 6.72万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10538129
• 关键词: Affect; Affinity; Antibodies; Apoptosis; Autoimmunity; Binding; Binding Proteins; Binding Sites; Biological Process; Biological Products; Breathing; Cells; Characteristics; Chimeric Proteins; Chromatin Structure; DNA; DNA Binding; DNA Binding Domain; DNA-Protein Interaction; Diabetes Mellitus; Disease; Engineering; Enhancers; Functional disorder; Future; Gene Expression; Gene Expression Regulation; Gene Silencing; Generations; Genes; Genetic Transcription; Genome; Genomic DNA; Genomics; Homeostasis; Investigation; Lead; Libraries; Link; Malignant Neoplasms; Mediating; Nature; Oncogenic; Permeability; Pharmacology; Phenotype; Preparation; Process; Promoter Regions; Property; Proteins; Proto-Oncogene Proteins c-myc; RNA; Research; Role; Route; Site; Solubility; Specificity; Structure; Technology; Testing; Therapeutic; antagonist; base; clinically relevant; cofactor; design; dimer; drug discovery; effective therapy; empowered; innovation; lead optimization; mimetics; nervous system disorder; novel strategies; programs; promoter; rational design; recruit; small molecule; success; synthetic construct; targeted treatment; therapeutically effective; tool; transcription factor

PROJECT ABSTRACT Transcription factors are key functional proteins that regulate gene expression through direct interaction with specific sequences of genomic DNA, thereby activating or repressing transcription. This highly conserved process controls many central biological functions including cellular homeostasis, differentiation, and apoptosis. Given this central role in regulating cell state and function, it is unsurprising that aberrant activity is implicated in many diseases including neurological disorders, autoimmunity, diabetes, and cancer. In particular, the master regulator MYC transcription factor is dysregulated in over half of all cancers, yet effective therapies targeting MYC-driven gene programs have yet to be developed. While approaches involving direct targeting of MYC with small molecules and antibodies or indirect targeting such as gene silencing have been explored, these are associated with major drawbacks including weak binding interactions or activity, poor pharmacological properties, and off-target effects. As an alternative approach, the Moellering lab recently developed a new class of hyperstable synthetic DNA-binding domains derived from the transcription factor MAX that orthogonally self- dimerize to sequence-specifically bind DNA with high affinity and specificity. These synthetic transcriptional regulators (STRs) competitively inhibit the DNA binding of native dimers of MYC/MAX or MAX/MAX and contest MYC-dependent gene expression. Building on this precedence, I hypothesize that the potency of these compounds would be increased by covalently ligating split monomeric STRs to produce pre-organized dimeric STRs. Additionally, I hypothesize that generating multi-functional STRs that recruit transcriptional machinery to directed genomic sites would provide further control to inhibit MYC-dependent phenotypes. The proposed research aims to explore these two potential routes for advancing the STR technology and evaluate their effects on MYC-dependent gene expression and phenotypes. Together, this tandem approach will establish a new class of potent MYC antagonists and provide mechanistic tools to study oncogenic gene expression in cells. Additionally, this modular design strategy to develop transcription factor mimetics will provide a framework to enable preparation of future constructs tailored towards other important transcription factors and genomic targets.

项目摘要 转录因子是关键的功能蛋白，通过与转录因子的直接相互作用来调节基因表达。 基因组DNA的特定序列，从而激活或抑制转录。这种高度保守的 该过程控制许多中心生物学功能，包括细胞内稳态、分化和凋亡。 考虑到这种在调节细胞状态和功能中的核心作用， 许多疾病，包括神经障碍、自身免疫、糖尿病和癌症。特别是，大师 调节MYC转录因子在超过一半的癌症中失调，但有效的治疗靶向 MYC驱动的基因程序尚未开发。虽然涉及直接针对MYC的方法， 已经探索了小分子和抗体或间接靶向如基因沉默， 与主要缺点相关，包括结合相互作用或活性弱、药理学较差 属性和脱靶效应。作为替代方法，Moellering实验室最近开发了一种新的类 超稳定的合成DNA结合域来自转录因子MAX， 二聚化以高亲和力和特异性地序列特异性结合DNA。这些合成的转录 竞争性抑制MYC/MAX或MAX/MAX的天然二聚体的DNA结合， MYC依赖性基因表达。基于这一优先顺序，我假设这些药物的效力 通过共价连接分裂的单体STR以产生预组织的二聚体， 可疑交易报告。此外，我假设产生多功能STR，招募转录机器， 定向基因组位点将提供进一步的控制以抑制MYC依赖性表型。拟议 研究的目的是探索这两种可能的发展STR技术的途径，并评估它们的效果 MYC依赖的基因表达和表型。总之，这种串联方法将建立一个新的类 有效的MYC拮抗剂，并提供机制的工具，研究致癌基因在细胞中的表达。 此外，这种开发转录因子模拟物的模块化设计策略将提供一个框架， 能够制备针对其他重要转录因子和基因组的未来构建体 目标的