Investigation of the design, structure and mechanism of Mena protein interaction inhibitors

Mena蛋白相互作用抑制剂的设计、结构和机制研究

基本信息

批准号：
10408668
负责人：
Jackson Halpin
金额：
$ 6.98万
依托单位：
MASSACHUSETTS INSTITUTE OF TECHNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-06-01 至 2023-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10408668
关键词：
Actins Affinity Behavior Binding Biological Assay Biology Biophysics Blood Circulation Body part C-terminal Cancer Patient Cell Culture Techniques Cells Cessation of life Characteristics Collaborations Communication Complex Computer Models Crystallography Data Development Distal Distant Educational process of instructing Engineering Environment Epitopes Evaluation Family Goals Hand Human Institutes Invaded Investigation Laboratories Lead Libraries Link Malignant Neoplasms Mammalian Cell Mediating Mentors Metastatic to Methods Modeling Modernization Molecular Mutation Neoplasm Metastasis Pathway interactions Peptides Permeability Primary Neoplasm Process Proline Property Protein Engineering Protein Family Protein Isoforms Proteins Proteome RNA Splicing Regulation Regulator Genes Research Research Methodology Research Training Role Site Specificity Structure Techniques Testing Therapeutic Tissues Training Transfection Travel Variant X-Ray Crystallography anticancer research base biophysical techniques cancer cell cancer invasiveness cell motility design experimental study genetic regulatory protein high throughput screening inhibitor insight knock-down member metastatic process neoplastic cell overexpression paralogous gene preference protein function protein protein interaction responsible research conduct scaffold screening skills small molecule inhibitor therapeutic evaluation tool tumor vasodilator-stimulated phosphoprotein

项目摘要

Project Summary/Abstract Cancer metastasis depends on coordinated cytoskeletal processes induced by a characteristic change in expression of specific motility and actin-regulatory genes. Mena, a member of the Ena/VASP family of actin regulatory proteins, is highly upregulated in invasive cancer cells. The Ena/VASP proteins localize to actin-based assemblies via their structurally similar EVH1 domains which bind to short linear motifs (SLMs) in other proteins. Mena is integral to motility pathways that are characteristic of invasive cancer cells. An invasion-associated splice variant of Mena, MenaINV, has far more potent effects on metastasis than Mena and is preferentially expressed in invasive cancer cells. However, determination of the precise mechanistic roles of Mena and MenaINV in metastatic processes has proven challenging. There is currently no molecular explanation for differences in the protein-protein interaction properties of Mena and its paralogs/isoforms. Designed peptide/mini-protein binders can reveal molecular determinants of binding specificity and inspire/inform the design of lead inhibitors. They can also be used to probe the function of proteins in their cellular context and with temporal control, providing a direct evaluation of therapeutic potential. The primary goal of this proposal is to uncover the molecular basis for differences in the protein-interaction properties of Mena and MenaINV, determine the molecular origin of the binding specificity of an existing mini-protein inhibitor, and use this information to design and test paralog- and isoform- selective, cell-permeable mini-protein inhibitors of Mena. This goal will be accomplished by applying an array of biophysical experiments such as NMR, SAXS, X-ray crystallography and binding assays to uncover the molecular origin of protein-interaction differences between Mena and its paralogs/isoforms. These experiments will reveal molecular determinants of inhibitor binding specificity and contribute to our understanding of metastasis by providing a molecular explanation for differences between Mena and MenaINV. The biophysical information gained from these experiments will then be incorporated into the design of paralog- and isoform- specific mini-protein inhibitors using cutting-edge protein design methods, including structure-based computation, focused library design, and high throughput screening techniques. In addition to training in a variety of new research methods, the training plan outlined here includes extensive development of scientific communication, responsible conduct of research, scientific networking, teaching, mentoring and management skills. The research and training will take place in the laboratory of Dr. Amy Keating, a highly interdisciplinary and collaborative group at the forefront of the protein-protein interaction and protein design fields. The Keating lab is embedded in the MIT Biology department and part of the Koch Institute for Integrative Cancer Research, providing an ideal environment for training, collaboration, and research.

项目概要/摘要癌症转移取决于由特征性变化引起的协调的细胞骨架过程特定运动和肌动蛋白调节基因的表达。 Mena，肌动蛋白 Ena/VASP 家族的成员调节蛋白在侵袭性癌细胞中高度上调。 Ena/VASP 蛋白定位于基于肌动蛋白的通过结构相似的 EVH1 结构域进行组装，该结构域与其他蛋白质中的短线性基序 (SLM) 结合。 Mena 是侵袭性癌细胞特征的运动途径的组成部分。与入侵相关 Mena 的剪接变体 MenaINV 对转移的影响比 Mena 强得多，并且优先在侵袭性癌细胞中表达。然而，确定 Mena 和事实证明，转移过程中的 MenaINV 具有挑战性。目前尚无分子解释 Mena 及其旁系同源物/亚型的蛋白质-蛋白质相互作用特性的差异。设计肽/微型蛋白结合物可以揭示结合特异性的分子决定因素并启发/告知先导抑制剂的设计。它们还可用于探测细胞环境中蛋白质的功能，并与时间控制，提供治疗潜力的直接评估。该提案的主要目标是揭示 Mena 和 MenaINV 蛋白质相互作用特性差异的分子基础，确定现有微型蛋白抑制剂的结合特异性的分子起源，并使用这些信息可用于设计和测试旁系同源物和异构体选择性、细胞可渗透的微型蛋白抑制剂梅纳。这一目标将通过应用一系列生物物理实验来实现，例如 NMR、SAXS、 X 射线晶体学和结合测定揭示蛋白质相互作用差异的分子起源 Mena 及其旁系同源物/亚型之间。这些实验将揭示抑制剂的分子决定因素结合特异性并通过提供分子解释来帮助我们理解转移 Mena 和 MenaINV 之间的差异。从这些实验中获得的生物物理信息将被使用尖端蛋白质将其纳入旁系同源物和亚型特异性微型蛋白质抑制剂的设计中设计方法，包括基于结构的计算、重点库设计和高通量筛选技术。除了各种新研究方法的培训之外，这里概述的培训计划还包括广泛发展科学传播、负责任的研究行为、科学网络、教学、指导和管理技能。研究和培训将在博士的实验室进行。 Amy Keating，一个高度跨学科和协作的团队，处于蛋白质-蛋白质相互作用的最前沿和蛋白质设计领域。基廷实验室隶属于麻省理工学院生物系，也是科赫实验室的一部分综合癌症研究所，为培训、合作和研究提供理想的环境。