Identifying Endogenous Retroviral Factors in Viral Lymphomagenesis

项目摘要/摘要 恶性肿瘤是艾滋病毒感染者(PLWH)死亡的主要原因。在这些恶性肿瘤中， 非霍奇金淋巴瘤(NHL)是影响PLWH的最常见的血液肿瘤。尽管 采用高效抗逆转录病毒疗法(ART)，PLWH患NHL的可能性是 那些没有感染艾滋病毒的人。爱泼斯坦-巴尔病毒(Epstein Barr Virus，EBV)是一种与淋巴瘤关系最为密切的致癌病毒 通常在PLWH中观察到。活动性艾滋病毒感染可导致艾滋病 未经治疗的人，众所周知，艾滋病会促进EBV+淋巴瘤的发展，称为艾滋病- 定义癌症(ADC)，宿主因素，包括内源性逆转录病毒对淋巴肿大的影响 ART治疗具有免疫功能的PLWH，仍有待探索。塞萨曼和尼克松实验室的工作 研究表明，EBV和HIV各自劫持宿主细胞信号通路，诱导其表达 内源性逆转录病毒，包括作为人类内源性逆转录病毒(HERV)。要确定 这些病毒在淋巴癌发生中的作用，我们将从体外淋巴瘤模型中产生数据来测试 EBV和HIV蛋白对内源性逆转录病毒癌蛋白表达的影响此外，我们还将 对来自PLWH的弥漫性大B细胞淋巴瘤(DLBCL)进行批量和单细胞测序，以确定 肿瘤微环境(TME)细胞群的变化。我们假设会有独一无二的 具有与调控失调相关的癌前转录特征的EBV感染B细胞群体 HERV表达，包括HERVK(HML2)Np9癌基因的表达。 在目标1中，我们将确定EBV潜伏期III LMP2A和细胞外HIV p17变异对 Np9癌基因的表达。我们将进行逐步的分子和细胞研究，以确定 外源性病毒感染对内源性逆转录病毒癌基因表达的翻译影响。我们 还将进行多组分析，以确定与转录相关的病毒和宿主因素 病毒介导的Np9癌基因表达机制。 在目标2中，我们将使用我们建立的新的单细胞管道来确定位点特异性内源性逆转录病毒 来自PLWH的DLBCL样本的转录。我们在尼克松实验室工作的初步数据强烈表明 我们将能够使用我们开发的管道来识别和量化这些Herv转录 来自RNA测序数据。我们将评估EBV阳性样本相对于EBV阳性样本的相对细胞数量 是EBV-并确定宿主基因、逆转录病毒和病毒转录本的差异。此外，我们打算 从更大的EBV+/-新鲜冰冻组织队列中进行批量RNA测序，验证这些单细胞发现 来自PLWH的DLBCL。

PROJECT SUMMARY/ABSTRACT Malignancy is the leading cause of mortality amongst people living with HIV (PLWH). Of these malignancies, Non-Hodgkin lymphoma (NHL) is the most common hematological neoplasm affecting PLWH. Despite the introduction of highly active antiretroviral therapy (ART), PLWH are 10-20 times more likely to develop NHL than those without HIV. Epstein Barr Virus (EBV) is an oncogenic virus that is associated with the lymphomas most commonly observed in PLWH. Active HIV infection can lead to Acquired Immunodeficiency Syndrome (AIDS) in untreated people, and it is known that AIDS promotes the development of EBV+ lymphomas, termed AIDS- defining cancers (ADCs), the effect of host factors, including endogenous retroviruses on lymphomagenesis in immunocompetent PLWH treated with ART, remains to be explored. Work in the Cesarman and Nixon labs has shown that EBV and HIV each hijack host cell signaling pathways upon infection to induce the expression of endogenous retroelements, including as human endogenous retroviruses (HERV). To determine the contributions of these viruses on lymphomagenesis, we will generate data from in vitro lymphoma models to test the effect of EBV and HIV proteins on the expression of endogenous retroviral oncoproteins. Additionally, we will perform bulk and single-cell sequencing of Diffuse Large B Cell Lymphoma (DLBCL) from PLWH to determine the changes in cell populations of the tumor microenvironment (TME). We hypothesize that there will be unique populations of EBV infected B cells with pre-malignant transcriptomic signatures that correlate with dysregulated HERV expression, including expression of the HERVK(HML2) Np9 oncogene. In Aim 1, we will determine the impact of EBV latency III LMP2A and extracellular HIV p17 variants on the expression of the Np9 oncogene. We will perform stepwise molecular and cellular studies to determine the translational impact of exogenous viral infection on the expression of this endogenous retroviral oncogene. We will additionally perform multi-omic analysis to identify viral and host factors relevant to the transcriptional machinery of virus-mediated Np9 oncogene expression. In Aim 2, we will use our novel established single cell pipeline to determine locus-specific endogenous retroviral transcription of DLBCL samples from PLWH. Preliminary data from our work in the Nixon lab strongly suggests that we will be able to use the pipeline that we have developed to identify and quantify these HERV transcripts from RNA-sequencing data. We will assess the relative cell populations of samples that are EBV+ to those that are EBV- and determine the differences in host gene, retroviral and viral transcripts. Additionally, we intend to validate these single cell findings with bulk RNA-sequencing from a larger cohort of EBV+/- fresh frozen tissue of DLBCL from PLWH.