Stapled Peptides for Protein Interaction Research and Therapeutic Targeting in Human Cancer

项目概要/摘要 失调的蛋白质相互作用有助于人类细胞的发育、维持和化学耐药性 癌症。致癌蛋白质的关键接触点通常由所谓的“凹槽螺旋”介导 相互作用，一种蛋白质的α螺旋子成分插入另一种蛋白质的表面凹槽 驱动致癌信号传导。例如，BCL-2 家族蛋白调节细胞之间的关键平衡。 生与死和癌细胞过度表达抗凋亡成员，该成员含有表面凹槽， 有效地捕获促凋亡成员的“杀手”螺旋，以强制细胞永生。我们已经插入了所有- 碳氢化合物伸展成天然α螺旋以恢复其形状、稳定性和生物活性，以便它们可以 既可以用作剖析致癌蛋白相互作用的强大化学工具，也可以用作原型疗法 给他们下药。在过去的 15 年里，我在设计钉合肽方面积累了特殊的专业知识， 在癌症研究和治疗中的多种应用。我已经生成了 NMR 结构的自旋标记类似物 分析，用于通过质谱法快速结合位点分析的光反应结构，半胱氨酸反应 用于共价靶向致癌蛋白的变体、用于结合亲和力定量的荧光衍生物和 细胞成像、用于体内成像的放射性标记构建体以及用于临床前的迭代优化类似物 测试和翻译。钉合肽的显着影响最好地体现在其识别能力上。 新的癌症靶点、机制和药物结合位点及其在成人临床试验中的进展 以及癌症复发的儿童。这一竞争性续订申请的目标是继续扩大 钉合肽创新的广度和深度，以支持关键的癌症研究项目 这些试剂用于蛋白质组学发现、结构测定、作用机制研究和治疗 发展。具体来说，我的目标是大力支持我的部门主任的 NCI 资助的 R35 研究计划， Loren Walensky 博士是一名化学生物学家和儿科肿瘤学家，专注于 BCL-2 的表征 通过中和线粒体凋亡途径来驱动人类癌症的家庭相互作用机制。 重新激活癌症中的细胞凋亡对于克服化疗耐药性和病理性α螺旋至关重要 BCL-2 家族的相互作用非常适合通过钉合肽进行询问。在运行装订 Walensky 实验室和丹娜—法伯癌症化学生物学项目的肽设计小组，我是 亲自负责开发和优化推动装订肽创新的化学，以及 创建高通量咨询、生产、纯化、定量和表征工作流程 我们的数十个内部和外部合作者也依赖它。作为一名研究专家，在 化学、癌症生物学和实验治疗学的交叉点，我致力于挖掘潜力 下一代钉合肽以增进我们对基本致癌机制的理解 创造癌症治疗的新疗法。

PROJECT SUMMARY / ABSTRACT Deregulated protein interactions contribute to the development, maintenance, and chemoresistance of human cancer. The critical contact points of cancer-causing proteins are often mediated by so-called “helix-in-groove” interactions, whereby an alpha-helical subcomponent of one protein inserts into the surface groove of another to drive oncogenic signaling. For example, BCL-2 family proteins regulate the critical balance between cellular life and death and cancer cells overexpress the anti-apoptotic members, which contain a surface groove that effectively traps the “killer” helix of pro-apoptotic members, to enforce cellular immortality. We have inserted all- hydrocarbon struts into natural alpha-helices to restore their shape, stability, and bioactivity so that they can be used as both powerful chemical tools to dissect oncogenic protein interactions and as prototype therapeutics to drug them. Over the last 15 years, I have developed special expertise in the design of stapled peptides for diverse applications in cancer research and treatment. I have generated spin-labeled analogs for NMR structural analyses, photoreactive constructs for rapid binding-site analysis by mass spectrometry, cysteine-reactive variants for covalent targeting of oncogenic proteins, fluorescent derivatives for binding affinity quantitation and cellular imaging, radiolabeled constructs for in vivo imaging, and iteratively-optimized analogs for preclinical testing and translation. The remarkable impact of stapled peptides is best reflected by their capacity to identify new cancer targets, mechanisms, and druggable binding sites and their advancement to clinical trials in adults and children with relapsed cancers. The goal of this competitive renewal application is to continue to expand the breadth and depth of stapled peptide innovation in support of critical cancer research programs that harness these reagents in proteomic discovery, structural determination, mechanism-of-action studies, and therapeutic development. Specifically, I aim to robustly support the NCI-funded R35 research program of my Unit Director, Dr. Loren Walensky, who as a chemical biologist and pediatric oncologist, focuses on characterizing the BCL-2 family interaction mechanisms that drive human cancer by neutralizing the mitochondrial apoptosis pathway. Reactivating apoptosis in cancer is essential to overcoming chemoresistance and the pathologic alpha-helical interactions of the BCL-2 family are ideally suited for interrogation by stapled peptides. In running the Stapled Peptide Design Group of the Walensky lab and Dana-Farber’s Program in Cancer Chemical Biology, I am personally responsible for developing and optimizing the chemistry that drives stapled peptide innovations, and creating a high-throughput consultation, production, purification, quantitation, and characterization workflow that is also relied upon by dozens of our internal and external collaborators. As a Research Specialist operating at the interface of chemistry, cancer biology, and experimental therapeutics, I am committed to mining the potential of next-generation stapled peptides to advance our understanding of fundamental oncogenic mechanisms and to create novel therapeutics for cancer treatment.