Analysis of the E7-Mediated Mechanism of MHC Class I Repression in HNSCC

E7 介导的 HNSCC I 类 MHC 抑制机制分析

• 批准号: 10219154
• 负责人: Elizabeth Gensterblum-Miller
• 金额: $ 3.94万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-01 至 2023-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10219154
• 关键词: 3-Dimensional; Acetylation; Address; Alanine; Antitumor Response; Architecture; Autoantigens; Binding; Binding Sites; Biological Assay; CD3 Antigens; CD8-Positive T-Lymphocytes; CD8B1 gene; CRISPR/Cas technology; Cell Line; Cells; Cessation of life; ChIP-seq; Chromatin; Chromatin Structure; Clinical Trials; Clustered Regularly Interspaced Short Palindromic Repeats; Coculture Techniques; Cytotoxic T-Lymphocytes; DNA; Data; Detection; Disease; Etiology; Flow Cytometry; Gene Expression Regulation; Genes; Genetic; Genetic Transcription; Goals; HLA-A gene; HPV-High Risk; Head and Neck Squamous Cell Carcinoma; Human Papillomavirus; Human papillomavirus 16; Human papillomavirus 18; Immune; Immune Evasion; Immunotherapy; Incidence; Interferon Type II; Knock-out; MHC Class I Genes; Major Histocompatibility Complex; Malignant Neoplasms; Mass Spectrum Analysis; Mediating; Modeling; Molecular; Oncogenic; Oncoproteins; Organoids; Pathogenicity; Pathway interactions; Patients; Peripheral Blood Mononuclear Cell; Pharmacologic Substance; Production; Proteins; RNA Polymerase II; Regulation; Regulatory Pathway; Repression; Scanning; Site; Stains; Surface; T-Cell Activation; T-Lymphocyte; Testing; Therapeutic; Transcriptional Regulation; Transfection; United States; Validation; Work; anti-tumor immune response; base; cancer type; genome-wide; genomic locus; human old age (65+); improved; inhibitor/antagonist; knockout gene; malignant oropharynx neoplasm; neoantigens; neoplastic cell; new therapeutic target; novel; novel therapeutic intervention; overexpression; prevent

Abstract Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer type in the world, and is responsible for over 8,000 deaths in the United States each year. The number of HNSCC cases caused by human papilloma virus (HPV) is on the rise, especially in patients 40-65 years old. HPV is currently associated with 80% of oropharyngeal cancers and 5-10% of HNSCC cases from other sub-sites. High-risk HPV strains, most commonly HPV16 and HPV18, cause the preponderance of HPV-positive HNSCC. Current data suggests that in HNSCC, HPV16 limits immune detection by preventing surface presentation of the major histocompatibility complex class I (MHC-I). In HPV-positive HNSCC, MHC-I surface presentation level is associated with the cytotoxic T cell-mediated anti-tumor immune response and thus the identification of strategies to prevent HPV-mediated MHC-I repression may have a significant therapeutic benefit for patients receiving immunotherapies. There some data that suggest E7 negatively regulates transcription of MHC-I constituent genes; however, the detailed mechanism of E7-mediated MHC-I repression is poorly characterized. Here, I have generated two E7-overexpressing HPV-negative HNSCC cell lines and confirmed that E7 protein downregulates transcription of MHC-I. We have developed systematic and logical approaches including CRISPR/CAS9 profiling that we propose to leverage to characterize the molecular mechanism(s) by which E7- represses MHC-I in these models, and HPV+ HNSCC models. Further, we have developed a 3D organoid-based co-culture assay in which these models are co-cultured with patient-matched peripheral blood mononuclear cells (PBMCs) to test the functional effects of de-repression of MHC-I expression. My central hypothesis is that identification of the molecular mechanisms by which HPV16_E7 diminish the expression of MHC-I will lead to the advancement of therapeutic strategies that enhance tumor cell recognition by activated T- cells. I will address this hypothesis through the following aims: 1) Detail the molecular mechanism(s) of MHC locus repression by HPV16_E7 in HNSCC, and 2) Qualify HPV16_E7-dependent MHC class I regulatory pathways in HNSCC. My long-term goal is to develop new therapeutic approaches that improve the overall survival of HPV+ patients, and in doing so, I hope to characterize the specific mechanisms by which HPV16_E7 can prevent MHC-I surface expression in HPV-positive HNSCC.

摘要 头颈部鳞状细胞癌(HNSCC)是世界上第六大最常见的癌症类型， 每年在美国造成超过8000人死亡。由HNSCC引起的HNSCC病例数量 人乳头瘤病毒(HPV)呈上升趋势，尤其是在40-65岁的患者中。HPV当前已关联 80%的口咽癌和5-10%的HNSCC病例来自其他亚区。高危HPV毒株， 最常见的是HPV16和HPV18，导致HPV阳性HNSCC的优势。目前的数据表明 在HNSCC中，HPV16通过阻止主要表型的出现来限制免疫检测 组织相容性复合体I类(MHC-I)。在HPV阳性的HNSCC中，MHC-I表面呈现水平为 与细胞毒性T细胞介导的抗肿瘤免疫反应相关，从而鉴定 预防HPV介导的MHC-I抑制的策略可能对患者有显著的治疗益处 接受免疫治疗。有一些数据表明，E7对MHC-I的转录具有负面调节作用 然而，E7介导的MHC-I抑制的详细机制尚不清楚。 在这里，我已经建立了两个过表达HPV阴性的HNSCC细胞系，并证实了E7蛋白 下调MHC-I的转录。我们制定了系统和合乎逻辑的方法，包括 我们建议利用CRISPR/Cas9图谱来表征E7-E7-的分子机制(S) 在这些模型和HPV+HNSCC模型中抑制MHC-I。此外，我们还开发了一种基于3D有机物的 将这些模型与患者匹配的外周血单核细胞共培养的共培养试验 (PBMCs)，以测试MHC-I表达下调的功能效应。我的中心假设是 HPV16_E7抑制MHC-I基因表达的分子机制研究 导致通过激活T细胞增强肿瘤细胞识别的治疗策略的进步 细胞。我将通过以下目的来解释这一假说：1)详细阐述MHC的分子机制(S) HPV16_E7在HNSCC中的位点抑制，以及2)HPV16_E7依赖的MHC I类调控 HNSCC中的路径。我的长期目标是开发新的治疗方法，以改善整体 HPV+患者的存活，在这样做的过程中，我希望描述HPV16_E7 可阻止HPV阳性HNSCC中MHC-I的表面表达。