Roles of LMP1 and MYC in EBV-induced B-cell tumors

LMP1 和 MYC 在 EBV 诱导的 B 细胞肿瘤中的作用

• 批准号: 10749776
• 负责人: Shannon Celeste Kenney
• 金额: $ 45.12万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2028-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10749776
• 关键词: B-Cell Lymphomas; B-Cell Neoplasm; B-Lymphocytes; Biological Models; Burkitt Lymphoma; Cell Culture Techniques; Cell Line; Cells; DNA Methylation; DNA Methylation Inhibition; DNMT3B gene; Development; EBNA2 protein; Epstein-Barr Virus Infections; Epstein-Barr Virus latency; Epstein-Barr Virus-Related Lymphoma; Gene Mutation; Genes; Genetic Transcription; Growth; HIV Infections; Hodgkin Disease; Human; Human Herpesvirus 4; Immune response; Immunocompetent; Immunocompromised Host; In Vitro; LMP1; Lymphoma; MYC gene; Mediating; Modeling; Mus; Oncogenic Viruses; Pathway interactions; Patients; Pharmaceutical Preparations; Phenotype; Proteins; RNA; Retroviral Vector; Retroviridae; Role; Structure of germinal center of lymph node; System; T cell response; T-Lymphocyte; TNFSF5 gene; Testing; Time; Tonsil; Umbilical Cord Blood; Viral; Viral Proteins; Virus Latency; anti-tumor immune response; experimental study; humanized mouse; immunogenic; immunosuppressed; in vitro Model; in vivo; in vivo Model; infected B cell; interleukin-21; large cell Diffuse non-Hodgkin' s lymphoma; mouse model; mutant; neoplastic cell; novel; overexpression; promoter; public health relevance; restoration; small molecule; stable cell line; transcription factor; transforming virus; tumor; viral RNA

PROJECT SUMMARY/ABSTRACT Epstein-Barr virus (EBV) is an important cause of human lymphomas in both immunocompetent and immunosuppressed humans, including Burkitt lymphomas (BLs), Hodgkin lymphomas (HLs) and diffuse large B cell lymphomas (DLBCLs). There are three different types of EBV latency (types I, II and III) that differ in the number of latent viral proteins expressed and transforming ability. Only type III viral latency (in which all 9 latent viral proteins are expressed) can transform primary human B cell in vitro, but because type III latency is highly immunogenic, EBV+ tumors with type III latency in humans are relatively rare and largely found in immunosuppressed hosts. EBV+ HLs and DLBCLs in humans commonly have type II latency (characterized by expression of EBNA1, LMP1 and LMP2A), while EBV+ BLs, which contain MYC translocations, have type I latency (in which EBNA1 is the only viral protein expressed). However, there is currently no in vivo or in vitro model available to study how EBV infection causes lymphomas with type I or type II latency, since this form of viral latency is not transforming in vitro and wild-type EBV-induced lymphomas in humanized mouse models inevitably support type III latency. EBNA2 transcriptionally activates each of the latent viral promoters used during type III latency. Using a newly constructed EBNA2-deleted EBV mutant (ΔEBNA2 EBV) made by our lab, we have developed a novel culture system that allows us to stably infect primary naïve B cells in vitro with this mutant, and to examine the effect of MYC over-expression. Our exciting preliminary results show that B cells infected with ΔEBNA2 EBV form DLBCL-like and HL-like tumors with type II viral latency at late time points in NSG mice, and that over-expressing the MYC gene (using a retroviral vector) in ΔEBNA2 EBV-infected B cells results in rapid onset of aggressive tumors that resemble human BLs and support type I EBV latency. In contrast, expression of MYC alone does not cause tumors in this model. As human BLs, DLBCLs and HLs are derived from germinal center (GC) B cells, in Aim 1, we will use this new model to examine the ability of ΔEBNA2 EBV- infected primary GC B cells (with or without MYC over-expression) to form lymphomas in NSG mice. In Aim 2, we will identify the specific EBV genes (and/or viral RNAs) required for the development of ΔEBNA2 EBV- induced tumors (with or without MYC) in naïve versus GC B cells. In Aim 3, we will define mechanisms by which MYC turns off LMP1 expression in ΔEBNA2 EBV-induced lymphomas and determine if small molecules that can restore LMP1 expression enhance the immune response to tumors in humanized mice. The proposed experiments will provide the first model system to define how types I and II EBV latency cause lymphomas in human GC B cells and to ask if restoration of LMP1 expression in BLs enhances the host immune response.

项目摘要/摘要 爱泼斯坦-巴尔病毒(Epstein-Barr Virus，EBV)是引起人类免疫活性淋巴瘤和非霍奇金淋巴瘤的重要原因。 免疫抑制的人类，包括Burkitt淋巴瘤(BLS)、霍奇金淋巴瘤(HLS)和弥漫性大B 细胞性淋巴瘤(DLBCL)。有三种不同类型的EBV潜伏期(类型I、II和III)在 潜伏病毒蛋白的表达数量和转化能力。只有III型病毒潜伏期(其中9个潜伏期 病毒蛋白表达)可以在体外转化原代人类B细胞，但因为III型潜伏期很高 在人类中，具有III型潜伏期的免疫原性EBV+肿瘤相对罕见，主要发现于 免疫抑制的宿主。人类的EBV+HLS和DLBCL通常有II型潜伏期(特征是 EBNA1、LMP1和LMP2a的表达)，而含有MYC易位的EBV+BLS为I型 潜伏期(其中EBNA1是唯一表达的病毒蛋白)。然而，目前还没有体内或体外的 可用于研究EBV感染如何导致I型或II型潜伏期淋巴瘤的模型，因为这种形式的 病毒潜伏期在体外没有转化，野生型EBV诱导的人源化小鼠模型中的淋巴瘤 不可避免地支持第三类延迟。EBNA2转录激活所用的每一种潜在病毒启动子 在III型潜伏期。使用本实验室新构建的EBNA2缺失EBV突变体(ΔEBNA2EBV)， 我们已经开发了一种新的培养系统，使我们能够在体外稳定地感染原代幼稚B细胞 突变株，并检测MYC过表达的影响。我们令人兴奋的初步结果表明，B细胞 感染Δ的EBNA2 EBV形成DLBCL样和HL样肿瘤，在晚期时间点有II型病毒潜伏 在ΔEBNA2 EB病毒感染的B细胞中过表达myc基因(使用逆转录病毒载体) 导致类似于人类BLS的侵袭性肿瘤的快速发病，并支持I型EBV潜伏期。相比之下， 在这个模型中，单独表达MYC并不会导致肿瘤。作为人的BLS，推导了DLBCL和HLS 在目标1中，我们将使用这个新的模型来检测ΔEBNA2 EBV-1的能力。 感染原代GC B细胞(有或不有MYC过表达)在NSG小鼠中形成淋巴瘤。在目标2中， 我们将确定发展ΔEBNA2 EB病毒所需的特定EB病毒基因(和/或病毒RNA)- 在幼稚细胞和GC B细胞中诱导肿瘤(有或没有MYC)。在目标3中，我们将定义通过哪些机制 MYC在ΔEBNA2EBV诱导的淋巴瘤中关闭LMP1的表达，并确定小分子是否可以 恢复LMP1的表达增强人源化小鼠对肿瘤的免疫应答。建议数 实验将提供第一个模型系统来定义I型和II型EBV潜伏期如何导致淋巴瘤 并询问在BLS中恢复LMP1表达是否能增强宿主的免疫反应。