Targeting the autophagy-lysosome system to block pancreatic cancer

靶向自噬-溶酶体系统来阻止胰腺癌

基本信息

批准号：
10590682
负责人：
Rushika Miriam Perera
金额：
$ 36.49万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
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：
2021-04-01 至 2026-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10590682
关键词：
Address Affinity Antigen Presentation Autophagocytosis Autophagosome BRCA1 gene Binding Biochemistry Biogenesis Cell Line Cell membrane Cell surface Cells Complex Coupled Endoplasmic Reticulum Endowment Ensure Epithelium Evolution Excision Genetic Goals Golgi Apparatus Grant Growth Human Immune Immune Evasion Immunofluorescence Immunologic In Vitro Location Lysosomes Major Histocompatibility Complex Malignant - descriptor Malignant Neoplasms Malignant neoplasm of pancreas Mediating Molecular Mus Neoplasm Metastasis Normal Cell Organelles Pancreas Pancreatic Ductal Adenocarcinoma Pancreatic Intraepithelial Neoplasia Pathway interactions Patients Peptides Post-Translational Protein Processing Proteins Proteomics Publications Recycling Role Route Sampling Specimen Stains Surface System T-Lymphocyte Testing Tissue Banks Tumor Tissue Ubiquitin Work cancer cell fitness genetic approach immune checkpoint blockade immunogenicity in vivo metabolic fitness mouse model novel pancreatic ductal adenocarcinoma cell pancreatic ductal adenocarcinoma model peptide I protein degradation receptor trafficking tumor ubiquitin ligase

项目摘要

PROJECT SUMMARY Cancer cells co-opt autophagy - an evolutionarily conserved cellular recycling pathway - to maintain metabolic fitness. Prior studies have shown that Pancreatic Ductal Adenocarcinoma (PDA) up-regulates autophagy and lysosomes – acidic organelles where autophagic cargo is degraded – to dramatically increase the bulk breakdown and recycling of diverse intracellular substrates. An additional, less well understood function of autophagy is the selective removal of specific proteins in order to endow PDA cells with enhanced fitness. We now show that the autophagy-lysosome pathway selectively targets major histocompatibility complex class I (MHC-I) protein for degradation as a mechanism of immune evasion. Through affinity- based capture of intact lysosomes (LysoIP) from normal and PDA cell lines coupled with proteomics-based analysis, we identified MHC-I as a significantly enriched lysosomal substrate in PDA cells. Consistent with this finding, immuno-fluorescence staining demonstrates that, unlike normal cells where MHC-I localizes to the plasma membrane (PM), in PDA cell lines and primary patient PDA specimens, MHC-I is trapped inside autophagosomes and lysosomes. Importantly, tumor-specific suppression of autophagy is sufficient to 1) stabilize and re-localize MHC-I to the PM, 2) increase antigen presentation and 3) boost T cell mediated tumor killing in vitro and in vivo. Building on these findings the goal of this study is to determine the molecular mechanisms underlying aberrant MHC-I trafficking, and to identify targetable nodes that can be manipulated to restore MHC-I on the cell surface of PDA cells. Our revised application will leverage the combined power of biochemistry, genetics, organelle purification and proteomics, a novel mouse model and patient PDA samples to address the following specific aims: 1) determine how post-translational modifications of MHC-I cooperate with the autophagy machinery to facilitate capture by autophagosomes, 2) identify where along its trafficking route is MHC-I diverted to autophagosomes and 3) determine when during the course of PDA evolution does MHC-I dysregulation occur and can we exploit this information to establish more effective strategies to enhance antigen presentation and immune mediated tumor killing. In summary, our discovery of autophagy dependent degradation of MHC-I highlights an important new paradigm for immune evasion in PDA and potentially other cancers. Findings from our proposed studies will determine key molecular mechanisms underlying MHC-I dysregulation and establish new nodes that can be targeted to restore antigen presentation in PDA.

项目概要癌细胞选择自噬（一种进化上保守的细胞回收途径）来维持先前的研究表明胰腺导管腺癌 (PDA) 上调。自噬和溶酶体（自噬货物被降解的酸性细胞器）急剧增加另一个不太为人所知的现象是不同细胞内底物的大量分解和回收。自噬的功能是选择性去除特定蛋白质，从而赋予 PDA 细胞增强体质。我们现在表明自噬-溶酶体途径选择性地针对主要组织相容性复合物 I 类 (MHC-I) 蛋白通过亲和力降解，作为免疫逃避的机制。基于从正常细胞系和 PDA 细胞系中捕获完整溶酶体 (LysoIP) 并结合基于蛋白质组学的通过分析，我们确定 MHC-I 是 PDA 细胞中显着富集的溶酶体底物，与此一致。研究发现，免疫荧光染色表明，与正常细胞不同，MHC-I 定位于质膜 (PM)，在 PDA 细胞系和原发患者 PDA 标本中，MHC-I 被困在内部重要的是，肿瘤特异性的自噬抑制足以：1) 稳定 MHC-I 并将其重新定位到 PM，2) 增加抗原呈递，3) 增强 T 细胞介导的肿瘤基于这些发现，本研究的目标是确定体外和体内杀灭作用的分子。异常 MHC-I 贩运的潜在机制，并识别可被操纵的目标节点恢复 PDA 细胞表面的 MHC-I 我们修改后的应用程序将利用综合力量。生物化学、遗传学、细胞器纯化和蛋白质组学、新型小鼠模型和患者 PDA 样本解决以下具体目标：1）确定 MHC-I 的翻译后修饰如何合作利用自噬机制促进自噬体捕获，2) 确定其运输的位置 MHC-I 转移至自噬体的路线，并且 3) 确定 PDA 进化过程中何时发生 MHC-I 失调发生，我们能否利用这些信息来制定更有效的策略来增强抗原呈递和免疫介导的肿瘤杀伤。总之，我们对 MHC-I 的自噬依赖性降解的发现凸显了一个重要的新发现我们提出的研究结果将成为 PDA 和潜在其他癌症中免疫逃避的范例。确定 MHC-I 失调的关键分子机制并建立新的节点旨在恢复 PDA 中的抗原呈递。