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 失调的细胞内蛋白质相互作用介导病理信号事件的复杂网络， 驱动人类癌症。药物的发展，以调节大，平，和复杂的界面，致癌 信号蛋白仍然是一个艰巨的挑战，并激发了替代方法，以传统的小 分子发现尽管定义了分子握手的肽结构 蛋白质理想地适合于调节这样的信号传导事件，在蛋白质的背景下结构化肽 当与整体分离时通常失去其生物活性形状。我们以前开发了碳氢化合物钉 通过插入化学交联而重现β-螺旋相互作用基序的天然形状的肽。 钉合肽是结构稳定的，抗蛋白酶的，并且保留了与其生物活性结合的能力。 具有天然效力和特异性的靶点。出乎意料的是，选择的钉合β-螺旋肽也是细胞- 渗透性，打开了一扇全新的模式，解剖和潜在的麻醉病理 癌症中的蛋白质相互作用。然而，尽管十年来在创造新的研究工具方面取得了进展， 研究和靶向致病蛋白，结构肽作为一种新型治疗药物的真正前景 我们对两个基本问题的理解非常有限，这阻碍了癌症的平台：（1） 什么样的生物物理特性决定了钉合肽是否会被癌细胞吸收？(2)是什么 明确的分子机制的细胞输入和细胞内释放的细胞渗透钉肽？ 这些问题的答案不仅具有改造这些研究工具和原型的潜力， 治疗剂进入真正的癌症药物库，但也将提供关键的新见解， 囊泡运输的细胞过程。例如，某些癌细胞通过以下方式实现代谢永生： 通过胞饮输入途径上调蛋白质营养摄取--一种可以利用的机制 以实现基于结构化肽的癌症药物的治疗窗口。在这里，我提出了一个双管齐下的建议， 利用我们在钉合肽设计和应用方面的丰富经验，但存在分歧的研究计划 从我们传统的蛋白质靶向研究，而是专注于为什么和如何不同的钉肽 进入细胞内环境为了实现我们的目标，我们将（1）采用生物统计和计算 方法来收集什么生物物理参数赋予我们的装订库中的细胞迁移率 肽，和（2）进行全基因组的CRISPR为基础的筛选，以确定，然后审查这些细胞 这些组分交替地损害和增强钉合肽的输入途径。通过整合我们的 实验室的基础化学生物学致癌蛋白相互作用的建议生物统计， CRISPR筛选和癌细胞输入验证研究，我们希望在我们的理解中开辟新天地 如何利用钉合肽及其摄取机制来治疗癌症。