The Impact of Epstein Barr Virus Infection on the Immune Response in Pediatric Transplant Recipients

7.项目摘要/摘要 实体器官移植已成为治疗患有各种终末期器官的儿童的主要方法 疾病。然而，预防移植物排斥所需的免疫抑制在移植部位 受体患严重机会性病毒感染的风险增加，这可能对移植物产生有害影响 和病人的生存。在儿科移植受者中，与EB病毒(EBV)相关的并发症有 特别令人担忧的。EB病毒感染的临床表现复杂，可从 与移植后淋巴组织增生性相关的侵袭性B细胞淋巴瘤的无症状病毒血症 精神障碍(PTLD)。移植中的一个主要问题是理解为什么有些孩子可以控制 EBV感染，而其他人则出现严重的危及生命的并发症。因此，需要新的战略来 开发更个性化的诊断和治疗儿童移植受者感染的方法 带着EB病毒。我们假设EBV感染塑造了移植后的先天免疫和获得性免疫 儿童的反应，这些变化可以被利用来识别独特的基于免疫的签名， 促进功能性EBV免疫和移植物长期存活。为了检验这一假设，我们建议使用 作为CTOT-C-06《移植后淋巴增殖性疾病的生物标志物》的一部分收集的额外样本 儿童“(PI：Esquivel)，并分析CTOT-C-02的现有数据，”儿童的免疫发育“ 移植“(PI：Kirk)。我们已经产生了强有力的初步数据表明：1)EBV病毒血症是 常见于儿童同种异体移植受者移植后第一年；2)NK细胞和T细胞增加 在EBV PTLD淋巴瘤和EBV-B细胞淋巴瘤皮损中的比较；3)NK细胞的一个特殊亚群 表达NKG2A能够对EBV感染的细胞产生应答和杀伤作用；4)克隆扩增的EBV- 利用TCR和抗原结合部分中的共享序列的特定T细胞可以在 EBV感染者；5)EBV B细胞淋巴瘤患者分泌免疫调节细胞因子 这会影响宿主的免疫反应。在第一个目标中，我们挖掘先前在CTOT-C-中获得的数据。 02研究病毒感染和临床结局背景下的NK细胞表型和T细胞特征。 我们确定了NKG2ANK细胞、病毒载量与EBV感染控制和利用的关系 用质量细胞术(细胞仪飞行时间，CyTOF)获得EBV反应性NKG2a的完整表型 细胞。在第二个目标中，我们使用单细胞分析将T细胞受体的使用和效应表型联系起来 揭示与EBV免疫保护相关的EBV特异性T细胞的特征。在《目标3》中我们 解决为什么许多免疫抑制患者停止治疗的谜团 EBV PTLD的诊断维持了他们的同种异体移植，而不会回到免疫抑制状态。我们使用CyTOF 为了确定在没有免疫抑制的情况下从EBV PTLD中恢复是否有利于 产生IL-10的B调节细胞的发展。总而言之，这些研究将提供新的机制 深入了解EBV的免疫反应，并将有助于改进诊断、管理和 儿科移植受者EB病毒病的治疗。

7. Project Summary/Abstract Solid organ transplantation has become a leading therapy for children with a variety of end stage organ diseases. However, the immunosuppression required for prevention of graft rejection places transplant recipients at increased risk of serious opportunistic viral infections that can have deleterious effects on graft and patient survival. In pediatric transplant recipients, Epstein Barr virus (EBV)-associated complications are of particular concern. The clinical manifestations of EBV infection are complex and can range from asymptomatic viremia to aggressive B cell lymphomas associated with post-transplant lymphoproliferative disorder (PTLD). A major question in transplantation has been to understand why some children can control EBV infection while others develop serious life-threatening complications. Thus, new strategies are needed to develop more personalized approaches for diagnosis and treatment of pediatric transplant recipients infected with EBV. We hypothesize that EBV infection shapes the post-transplant innate and adaptive immune response in children and that these changes can be exploited to identify unique immune-based signatures that promote functional EBV immunity and long-term graft survival. To test this hypothesis we propose to utilize extra samples collected as part of CTOT-C-06, “Biomarkers for Post-Transplant Lymphoproliferative Disorders in Children” (PI:Esquivel) and to analyze existing data from CTOT-C-02, “Immune Development in Pediatric Transplantation” (PI:Kirk). We have generated strong preliminary data indicating that: 1) EBV viremia is common in the first year post-transplant in pediatric allograft recipients; 2) NK cells and T cells are increased in lesions of EBV+ PTLD lymphomas compared to EBV- B cell lymphomas; 3) a specific subset of NK cells that express NKG2A+ is capable of responding to, and killing, EBV-infected cells; 4) clonally expanded EBV- specific T cells that utilize TCR with shared sequences within the antigen-binding portion can be identified in EBV-infected individuals; and 5) patients with EBV+ B cell lymphomas secrete immunomodulatory cytokines that can influence the host immune response. In the first Aim we mine data previously obtained in CTOT-C- 02 to investigate NK cell phenotypes and T cell signatures in the context of viral infection and clinical outcome. We determine the relationship between NKG2A+ NK cells, viral load, and control of EBV infection and utilize mass cytometry (cytometry time of flight, CyTOF) to obtain a complete phenotype of EBV-reactive NKG2A+ cells. In the second Aim we use single cell assays to link T cell receptor usage and effector phenotype to reveal the signature of EBV-specific T cells associated with immune protection from EBV. In Aim 3 we address the enigma of why many patients who have had immunosuppression halted as a first line response to the diagnosis of EBV+PTLD maintain their allograft without returning to immunosuppression. We use CyTOF to determine whether recovery from EBV+ PTLD in the absence of immunosuppression favors the development of IL-10-producing B regulatory cells. Together, these studies will provide novel mechanistic insights into the immune response to EBV and will facilitate improvements in diagnosis, management and treatment of EBV disease in pediatric transplant recipients.