Developing Autophagy-Targeting Chimeras and Optimizing Cell Penetration of Large-Molecule Therapeutics

开发自噬靶向嵌合体并优化大分子治疗的细胞渗透

基本信息

批准号：
10558145
负责人：
Joshua A Kritzer
金额：
$ 57.9万
依托单位：
TUFTS UNIVERSITY MEDFORD
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-03-01 至 2028-02-29
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10558145
关键词：
Address Adopted Algorithm Design Area Autophagocytosis Binding Biological Assay CRISPR screen Cells Cellular biology Chimera organism Cytosol DNA Damage Data Degradation Pathway Disease Endosomes Genetic Human Kinetics Learning Malignant Neoplasms Measures Mediating Methods Modeling Molecular Evolution Penetration Protac Protein Family Proteins Series Structure Testing Therapeutic Time Work cell type chemotherapy design drug development drug discovery inhibition of autophagy inhibitor innovation novel protein degradation protein protein interaction research and development screening small molecule inhibitor stapled peptide

Address Adopted Algorithm Design Area Autophagocytosis Binding Biological Assay CRISPR screen Cells Cellular biology Chimera organism Cytosol DNA Damage Data Degradation Pathway Disease Endosomes Genetic Human Kinetics Learning Malignant Neoplasms Measures Mediating Methods Modeling Molecular Evolution Penetration Protac Protein Family Proteins Series Structure Testing Therapeutic Time Work cell type chemotherapy design drug development drug discovery inhibition of autophagy inhibitor innovation novel protein degradation protein protein interaction research and development screening small molecule inhibitor stapled peptide

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项目摘要

The Kritzer lab focuses on inhibiting protein-protein interactions involved in autophagy. Autophagy is a protein degradation pathway that is active in all human cells, and inhibiting autophagy shows promise as a therapy for late-stage cancers especially in combination with DNA-damaging chemotherapies. Autophagy research and drug development currently rely on compounds that inhibit autophagy indirectly. Better, more specific inhibitors of autophagy would be broadly adopted. A large amount of genetics and cell biology work supports that inhibiting the LC3/GABARAP family of proteins can block autophagy selectively. In one series of projects, we will develop novel stapled peptide and small molecule inhibitors of LC3/GABARAP and evaluate them in models of late-stage cancers and other diseases. Because of our expertise in compounds that bind LC3/GABARAP proteins, we also propose to evaluate related compounds as autophagy-targeting chimeras (AUTACs). These compounds could be used to selectively degrade any proteins in the cell, similar to proteolysis-targeting chimeras (PROTACs) but potentially more versatile and easier to develop. Based on strong preliminary data that validate the AUTAC concept, we will develop novel AUTACs and demonstrate their ability to degrade endogenous proteins, unlocking a broad new area for drug development in targeted protein degradation. Over the course of developing stapled peptides as autophagy modulators, the Kritzer lab encountered a common problem in the field: how to measure the amount that actually reaches the cytosol. In an independent series of projects, the Kritzer lab has developed novel assays that quantitate the cytosolic penetration of large-molecule therapeutics. In this proposal, we describe new opportunities to address challenging problems in drug development for large-molecule therapeutics. We describe new methods to measure penetration to different cellular compartments in any cell type, including primary cells. We also describe a molecular evolution approach to develop a new “turn-on” assay that measures the real-time kinetics of cytosolic penetration. We describe pooled CRISPR screens to reveal the cellular components that mediate endosomal escape. Finally, we describe a novel screening platform that will allow us to screen thousands to millions of molecules at a time for those that are most cell-penetrant; the new screen will be incorporated into a design-test-learn cycle to produce data-driven design algorithms for cytosol-penetrant molecules. All together, these data will represent a huge leap in our understanding of structure-penetration relationships for several classes of large-molecule therapeutics.

Kritzer Lab的重点是抑制自噬所涉及的蛋白质蛋白质相互作用。自噬是一种蛋白质在所有人类细胞中都活跃的降解途径，并且抑制自噬显示出有望作为一种治疗晚期癌症，尤其是与DNA受损化学疗法的结合。自噬研究和目前，药物开发依赖于间接抑制自噬的化合物。更好，更具体的抑制剂将广泛采用自噬。大量遗传学和细胞生物学工作支持抑制LC3/Gabarap蛋白质家族可以选择性地阻断自噬。在一个系列项目中，我们将开发出新的分阶段肽和LC3/gabarap的小分子抑制剂，并将其评估后期癌症和其他疾病的模型。由于我们在结合LC3/gabarap蛋白的化合物方面具有专业知识，我们还建议评估相关化合物作为靶向自噬的嵌合体（Autacs）。这些化合物可用于选择性降解细胞中的任何蛋白质，类似于靶向蛋白水解的嵌合体（Protac），但可能更多多才多艺，更容易开发。基于验证AUTAC概念的强大初步数据，我们将开发新颖的自动，并证明其降解内源性蛋白的能力，解锁靶向蛋白质降解中药物开发的广泛领域。在开发阶段肽作为自噬调节剂的过程中，Kritzer实验室遇到了现场的常见问题：如何测量实际达到细胞质的数量。在独立的项目系列，Kritzer Lab开发了定量胞质的新颖测定法大分子疗法的渗透。在此提案中，我们描述了解决的新机会大分子疗法的药物开发中的挑战性问题。我们将新方法描述为测量任何细胞类型（包括原代细胞）中不同细胞隔室的渗透。我们也是描述一种分子进化方法，以开发一种新的“转到”测定法，以实时测量胞质渗透动力学。我们描述了合并的CRISPR屏幕以揭示细胞组件媒体内体逃脱。最后，我们描述了一个新颖的筛选平台，该平台将使我们能够筛选对于大多数细胞渗透剂的人来说，数千至数百万分子；新屏幕将是纳入设计测试测试周期，以产生数据驱动的设计算法分子。总之，这些数据将代表我们对结构渗透的理解的巨大飞跃几类大分子疗法的关系。