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：
10212065
负责人：
Rushika Miriam Perera
金额：
$ 38.64万
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/10212065
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 Ensure Epithelial 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 Molecular Genetics 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 base 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.

项目摘要癌细胞选择自噬 - 一种进化配置的细胞回收途径 - 维持代谢健身。先前的研究表明，胰腺导管腺癌（PDA）上调自噬和溶酶体 - 自噬货物降解的酸性细胞器 - 显着增加潜水细胞内底物的大量分解和回收。另一个不太了解的自噬的功能是选择性去除特定蛋白质，以便将PDA细胞赋予增强的健身。现在，我们表明自噬溶质体途径有选择地靶向主要的组织相容性复杂的I类（MHC-I）蛋白作为免疫抗性的机理降解。通过亲和力 - 基于正常和PDA细胞系的基于基于蛋白质组学的正常和PDA细胞系的完整溶酶体（lysoip）捕获分析，我们将MHC-I确定为PDA细胞中的显着富集溶酶体底物。与此一致发现，免疫荧光染色表明，与正常细胞不同，MHC-I本地化在质膜（PM），在PDA细胞系和主要患者PDA标本中，MHC-I被困在内部自噬体和溶酶体。重要的是，自噬的肿瘤特异性抑制足以达到1）将MHC-1稳定并重新定位到PM，2）增加抗原表现，3）增强T细胞介导的肿瘤在体外和体内杀死。在这些发现的基础上，这项研究的目的是确定分子 MHC-I贩运的基础机制，并确定可以操纵的目标节点恢复PDA细胞细胞表面上的MHC-I。我们修订的应用将利用生物化学，遗传学，细胞器纯化和蛋白质组学，一种新型的小鼠模型和患者PDA样品解决以下具体目的：1）确定MHC-I翻译后修改如何配合借助自噬的机械促进自噬体捕获，2）确定贩运的位置路由是MHC-I转移到自噬体，3）确定何时在PDA进化过程中 MHC-I失调发生，我们可以利用这些信息来建立更有效的策略以增强抗原表现和免疫介导的肿瘤杀死。总而言之，我们发现MHC-I自噬依赖性降解的发现突出了一个重要的新 PDA和其他癌症中免疫进化的范例。我们提出的研究的发现将确定MHC-1失调的关键分子机制，并建立可以是的新节点旨在恢复PDA中的抗原表现。