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Function of Wdfy4 in cross-presentation and immunity

Wdfy4在交叉呈递和免疫中的功能

基本信息

批准号：
10430144
负责人：
Kenneth M Murphy
金额：
$ 50.73万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-07-01 至 2024-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10430144
关键词：
Alloantigen Antibody Response Antigen-Presenting Cells Antigens Area Autoimmune B-Lymphocytes Bacteria Biology CD8-Positive T-Lymphocytes CRISPR screen Cancer Patient Cells Chimeric Proteins Clinical Clinical assessments Cross Presentation Data Defect Dendritic Cells Development Diagnostic Dissection Effectiveness Electron Microscopy Electrons Epitopes Failure Genes Genetic Genetic Transcription Goals Human Immune response Immunity Immunofluorescence Microscopy Immunoprecipitation Immunotherapy In Vitro Knockout Mice Label Lead Listeria monocytogenes Literature Location Lupus Malignant Neoplasms Mass Spectrum Analysis Methods Modeling Molecular Mus Parasites Pathway interactions Patients Physiological Process Property Proteins Role T cell response T-Lymphocyte Testing Therapeutic Toxoplasma gondii Translations Tumor Antigens Tumor Immunity Tweens Virus Virus Diseases Work adaptive immune response antigen-specific T cells cancer immunotherapy cancer therapy cancer type checkpoint inhibition dendritic cell vaccination genome wide association study immune checkpoint blockade immunogenic immunogenicity improved in vivo inhibitor innovation insight monocyte mouse model neoantigens novel response transcription factor tumor virtual

项目摘要

ABSTRACT Checkpoint blockade immunotherapy relies on releasing tumor-specific T cells from normal inhibitory control, and has significantly impacted treatment for several types of cancer. However, response rates in patients treated with checkpoint blockade still need to be improved. Non-responsiveness to checkpoint blockade could result from many causes, including failure of dendritic cell priming of CD8 T cells. Indeed, checkpoint blockade has recently been shown to be dependent on the cDC1 subset of dendritic cells for its effectiveness. The cDC1 lineage of dendritic cells is the major cross-presenting cell that primes CD8 T cells and has been the subject of our recent molecular and developmental analysis. We were the first to identify that the cDC1 lineage requires the BATF3 transcription factor and is critical in vivo for tumor rejection. Cross-presentation is central to the ability of cDC1 to prime tumor-specific DC8 T cells, but this process has remained poorly understood. We have undertaken a molecular dissection of the mechanisms of cross-presentation in cDC1. Our ra- tionale is that understanding cross-presentation at a basic level could be used to improve treatment of can- cer patients by reducing non-responsiveness to checkpoint blockade. In addition, this could potentially be used to improve dendritic cell vaccination approaches in cancer. This application builds on our discovery of the first gene that is absolutely required for cDC1 cross-presentation in vivo and is critical for anti-tumor re- sponses. We identified Wdfy4 in a CRISPR/Cas9 screen in primary cDC1 as required for cross-presentation and we have already developed the Wdfy4-/- mouse model which is the basis for the current application. Our preliminary data show that Wdfy4-/- mice have a profound defect in cross-presentation, lacking the ability to prime CD8 T cells against viruses and tumors. We propose to use this model to determine the full scope of Wdfy4's function in vivo and to determine the mechanism by which WDFY4 supports cross-presentation in cDC1. Further, we will determine the molecular and cellular mechanism for this function of the WDFY4 protein. First, we will determine the intracellular location of WDFY4 in DCs, by using immunofluorescence microscopy of epitope tagged WDFY4 proteins, by localizing WDFY4 using APEX2 fusion proteins and electron microsco- py, and by localizing endogenous WDFY4 in primary human and mouse cDC1. Second we will identify WDFY4 interacting partners that cooperate in cross-presentation in cDC1 by testing whether Clec9a or Dec205 interact with WDFY4, by identifying WDFY4 interacting proteins by immunoprecipitation and mass-spectrometry analy- sis, and by proximity labeling, and by testing if these interacting proteins are required for cross-presentation.

摘要