Biophysical and Mechanistic Determinants for Cancer Cell Import of Hydrocarbon-Stapled Peptides

癌细胞输入碳氢化合物肽的生物物理和机制决定因素

• 批准号: 9178990
• 负责人: Loren David Walensky
• 金额: $ 19.03万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-02 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9178990
• 关键词: Address; Affinity; Algorithms; Antineoplastic Agents; BCL1 Oncogene; BCL2 gene; BCL2L11 gene; Biological; Biology; Biostatistical Methods; CRISPR screen; CRISPR/Cas technology; Candidate Disease Gene; Cell physiology; Cells; Chemicals; Circular Dichroism; Complex; Computing Methodologies; Data Set; Disease; EZH2 gene; Environment; Event; Exhibits; Foundations; Gene Deletion; Glean; Goals; High Pressure Liquid Chromatography; Human; Hydrocarbons; KRAS2 gene; Laboratories; Libraries; Malignant Neoplasms; Measurement; Measures; Mediating; Metabolic; Methods; Modality; Molecular; Nutrient; Oncogenic; Pathologic; Pathway interactions; Penetrance; Peptide Hydrolases; Peptide Library; Peptides; Pharmaceutical Preparations; Pinocytosis; Population; Principal Component Analysis; Property; Protein Family; Proteins; Research; Resistance; Scanning; Screening for cancer; Shapes; Signal Transduction; Signaling Protein; Son of Sevenless Proteins; Specificity; Statistical Data Interpretation; Statistical Methods; Targeted Research; Therapeutic; Time; Translating; Vesicle Transport Pathway; base; biophysical properties; cancer cell; cancer therapy; crosslink; cytotoxicity; design; drug development; experience; genome-wide; genome-wide analysis; insight; interdisciplinary approach; novel therapeutics; peptide drug; peptide structure; prototype; small molecule; therapeutic target; tool; uptake; validation studies

PROJECT SUMMARY Deregulated intracellular protein interactions mediate a complex network of pathologic signaling events that drive human cancer. The development of drugs to modulate the large, flat, and complex interfaces of oncogenic signaling proteins remains a formidable challenge and has inspired alternative approaches to traditional small molecule discovery. Although the very peptide structures that define the molecular handshakes between proteins are ideally suited to modulate such signaling events, structured peptides in the context of a protein typically lose their bioactive shape when isolated from the whole. We previously developed hydrocarbon-stapled peptides that recapitulate the natural shape of -helical interaction motifs by insertion of chemical crosslinks. Stapled peptides are structurally-stable, protease-resistant, and retain the capacity to engage their biological targets with natural potency and specificity. Unexpectedly, select stapled -helical peptides are also cell- permeable, opening the door to an entirely new modality for dissecting and potentially drugging pathologic protein interactions in cancer. However, despite a decade of progress in the creation of new research tools to investigate and target disease-causing proteins, the true promise of structured peptides as a novel therapeutic platform for cancer has been hampered by our very limited understanding of two fundamental questions: (1) What biophysical properties dictate whether a stapled peptide will be taken up by a cancer cell? (2) What is the explicit molecular mechanism of cellular import and intracellular release for cell-permeable stapled peptides? The answers to these questions not only carry the potential to transform these research tools and prototype therapeutics into an arsenal of bona fide cancer drugs, but will also provide critical new insight into the essential cellular process of vesicle transport. For example, certain cancer cells enforce metabolic immortality by upregulating protein nutrient uptake via the pinocytosis import pathway – a mechanism that could be leveraged to achieve a therapeutic window for structured peptide-based cancer drugs. Here, I propose a two-pronged research plan that harnesses our deep experience with stapled peptide design and application, but diverges from our traditional protein targeting research to instead focus on the why and how distinct stapled peptides access the intracellular environment. To achieve our goals, we will (1) employ biostatistical and computational methods to glean what biophysical parameters confer cellular penetrance among our libraries of stapled peptides, and (2) perform a genome-wide CRISPR-based screen to identify and then vet those cellular components that alternatively impair and enhance the import pathway for stapled peptides. By integrating our laboratory’s foundation in the chemical biology of oncogenic protein interactions with the proposed biostatistical, CRISPR screening, and cancer cell import validation studies, we hope to break new ground in our understanding of just how stapled peptides and their uptake mechanisms can be harnessed for therapeutic benefit in cancer.

项目摘要 放松管制的细胞内蛋白相互作用介导了一个复杂的病理信号事件网络， 推动人类癌。药物的开发以调节致癌的大型，平坦且复杂的界面 信号蛋白仍然是一个巨大的挑战，并激发了传统小型的替代方法 分子发现。虽然定义分子握手的肽结构 蛋白质非常适合调节此类信号事件，在蛋白质的背景下结构化胡椒粉 与整体隔离时，通常会失去其生物活性形状。我们以前开发了碳氢化合物 通过插入化学交联的肽来概括螺旋相互作用基序的自然形状的肽。 固定肽具有结构稳定，抗蛋白酶，并保留吸引其生物学的能力 具有自然效力和特异性的目标。出乎意料的是，精选的上演螺旋辣椒也是细胞 - 渗透性，为剖析和潜在吸毒病理学的全新方式打开了大门 癌症中的蛋白质相互作用。然而，尽管在创建新的研究工具方面取得了十年的进步 调查和靶向引起疾病的蛋白质，这是结构化胡椒粉作为一种新型疗法的真正希望 我们对两个基本问题的非常有限的理解使癌症平台受到了阻碍：（1） 哪些生物物理特性决定了癌细胞是否将钉蛋白肽取代？ （2）什么是 细胞进口和细胞内释放的明确分子机制，可用于细胞可渗透的固定肽？ 这些问题的答案不仅具有改变这些研究工具和原型的潜力 治疗药物为真正的癌症药物库，但也将为重要的新见解提供重要的新见解。 囊泡转运的细胞过程。例如，某些癌细胞通过 通过皮细胞增多症进口途径上调蛋白质养分的摄取 - 一种可以利用的机制 为了实现结构化肽癌药物的治疗窗口。在这里，我提出了一个两管齐的 利用我们在上演肽设计和应用方面的深厚经验的研究计划，但分歧 从我们传统的蛋白质靶向研究到关注为什么和如何不同的滞留肽 访问细胞内环境。为了实现我们的目标，我们将（1）员工生物统计学和计算 收集我们钉书库中的生物物理参数会议的细胞渗透的方法 肽和（2）执行基于基因组CRISPR的屏幕以识别并审查那些细胞 会损害并增强上演肽的进口途径的组件。通过整合我们的 实验室与所提出的生物统计学的致癌蛋白相互作用的化学生物学基础 CRISPR筛查和癌细胞进口验证研究，我们希望在我们的理解中脱颖而出 可以利用固定肽及其吸收机制的方式来利用癌症的治疗益处。