Understanding the immune response changes to clinical interventions for Epstein-Barr virus infection prior to lymphoma development in children after organ transplants (UNEARTH)

摘要/概要 在EB病毒相关的癌症中，移植后淋巴组织增生性疾病（PTLD）是一种罕见的 但却是小儿实体器官移植的主要并发症。许多儿童是EB病毒血清阴性的时间 SOT，导致在强烈免疫抑制下的同种异体移植物原发性EBV感染， 慢性高病毒载量（CHVL）状态或PTLD的机会。纵向外周血EBV DNA核酸 检测（NAT）并没有改善PTLD发生的个体预测，可能是由于不同的SOT接受者 免疫反应。此外，这些患者接受EBV DNA血症的临床干预， 原因不明的回应。我们的SOT，传染病和免疫学专业人员团队将带来 新的和互补的专门知识，以弥补这些知识差距。我们将进行纵向T细胞和NK细胞 在1390个样本中进行免疫功能测定，结合局部和中心EBV和环节病毒NAT， 在3家主要儿童医院进行278次SOT（肾、肝、心、肺或肠）后第一年的5个时间点。 我们将完成以下具体目标，比较胸部和腹部SOT受体与原发性 EBV感染或CHVL状态：1.评估T细胞的表型和功能特征 “衰竭”和与EBV感染结果和NK细胞谱相关。假设：SOT接受者 发展CHVL状态显示出明显表型记忆分化和耗尽的CD 8+和CD 4 + T细胞 这些细胞分布受不同的炎症回路调节。我们将通过执行多项任务来实现这一目标- 用于表征T细胞表型和功能的光谱流式细胞术，以及Meso Scale Discovery平台 为了评估不同的病毒控制相关的血浆细胞因子/趋化因子，在初始复制阶段， 扩张、进展、CHVL或恢复状态。2.为了前瞻性地定义数量、表型和 NK细胞的功能状态，并与EBV感染结果相关。假设：NK细胞活化 与原发性感染一致，与清除率呈正相关，与持续性EBV呈负相关 复制的NK细胞功能障碍将在CHVL患者中发展，这些患者处于PTLD的最高风险中。我们将 利用我们已建立的多光谱流式细胞术面板，分析原发性EBV感染患者 并回答与激活状态、NK受体库和功能能力相关的问题。 3.确定外周血扭体病毒（TTV）DNA载量与EBV结局的相关性， 和NK细胞谱。假设：TTV负荷随着临床免疫抑制的减少而减少， EBV清除。我们将使用两种病毒的纵向全血NAT检测来实现这一目标， 共同的时间点，集中进行，以尽量减少实验室变异性。通过研究，我们将知道T和NK 在多种临床情况下对EBV的免疫应答。我们希望找到关键的免疫机制， 将预测尽管临床干预，但EBV清除不良或延迟，这可能导致新的翻译， 免疫治疗方法来预防PTLD或告知其他人群中的EBV肿瘤发生。

Abstract/Summary Among the Epstein-Barr virus associated cancers is post-transplant lymphoproliferative disease (PTLD), a rare but major complication of pediatric solid organ transplants (SOT). Many children are EBV-seronegative at time of SOT, leading to primary EBV infection from the allograft under intense immunosuppression, and a higher chance of a chronic high viral load (CHVL) state or PTLD. Longitudinal peripheral blood EBV DNA nucleic acid testing (NAT) has not improved the individual prediction of PTLD occurrence, likely due to variable SOT recipient immune responses. Further, these patients receive clinical interventions for EBV DNAemia, with incomplete responses for unknown reasons. Our team of SOT, infectious disease and immunology professionals will bring new and complimentary expertise to close these knowledge gaps. We will perform longitudinal T and NK cell immune function assays in conjunction with local and central EBV and anellovirus NAT in 1390 samples across 5 time points in the first year after 278 SOT (kidney, liver, heart, lung or intestine) at 3 major children's hospitals. We will accomplish the following Specific Aims, comparing thoracic and abdominal SOT recipients with primary EBV infection or CHVL state: 1. Assess the prospective phenotypic and functional features of T cell “exhaustion” and correlate with EBV infection outcomes and NK cell profile. Hypothesis: SOT recipients' that develop CHVL state display distinct phenotypic memory differentiation and exhausted CD8+ and CD4+ T cell profiles that are regulated by distinct inflammatory circuits. We will accomplish this aim by performing multi- spectral flow cytometry to characterize T cell phenotype and function, as well as Meso Scale Discovery platform to assess distinct viral control-relevant plasma cytokines/chemokines, during the phases of initial replication, expansion, progression, CHVL or recovery states. 2. To prospectively define the number, phenotype, and functional status of NK cells, and correlate with EBV infection outcomes. Hypothesis: NK cell activation will coincide with primary infection, and will correlate positively with clearance vs. negatively with persistent EBV replication. NK cell dysfunction will develop in patients with CHVL, who are at highest risk of PTLD. We will leverage our established multi-spectral flow cytometry panel and analyze patients with primary EBV infection after SOT and answer questions related to the activation status, NK receptor repertoire, and functional capacity. 3. Determine the association of peripheral blood torquetenovirus (TTV) DNA loads to EBV outcomes, T and NK cell profiles. Hypothesis: TTV loads reduce with clinical reductions in immunosuppression and predict EBV clearance. We will accomplish this aim using longitudinal whole blood NAT assays for both viruses at common time points, performed centrally to minimize lab variability. By study end, we will know the T and NK immune responses to EBV across multiple clinical situations. We expect to find key immune mechanisms that will predict poor or delayed EBV clearance despite clinical interventions, which may lead to new translational immunotherapy approaches to prevent PTLD, or inform EBV oncogenesis in other populations.