Towards a Mouse Model of Classical Hodgkin's Disease and PTLD

描述(申请人提供)：霍奇金氏病(HD)是西方世界最常见的淋巴瘤，恶性细胞是所谓的霍奇金和里德-斯特恩伯格(HRS)细胞，仅占淋巴瘤肿块的几个百分点。HRS细胞常被EB病毒(EBV)感染，表达EBV蛋白LMP1和LMP2a，分别部分模仿CD40活性辅受体和B细胞抗原受体(BCR)。以前的工作已经发现体细胞突变的B细胞，通常携带突变的BCR表达，作为HRS的祖细胞，导致了HD发病的场景，其中HRS细胞来自于LMP2A和LMP1表达挽救的凋亡前GC B细胞。尽管HRS细胞起源于B细胞，但它在很大程度上失去了B细胞特异的基因表达程序，并获得了T和/或髓系造血系典型基因的表达。我们假设，这种谱系不忠是由某些已知在HRS细胞中表达的转录因子(TF)干扰B细胞特异性基因表达程序所决定的，即Id2、ABF1和Notch1。再加上促进细胞存活和增殖的蛋白质的结构性表达，如核因子-kB家族的Tf、c-jun和Tf，最终可能会将GC B细胞重新编程为HRS细胞。在这一一般情况下，EBV感染是HD的主要初始转化事件，确实已知导致B细胞中Tf-kB、AP1、Id2和ABF1的上调。根据这一普遍假设，我们开发了一种策略，使用Cre介导的条件基因打靶将候选基因的表达定向到GC B细胞中。使用并进一步改进这些工具，我们计划分析在GC B细胞中单独和联合诱导各种EBV衍生蛋白和TF的表达是否会导致这些细胞的反式分化和转化，就像在HD的HRS细胞中看到的那样。我们的最终目标是建立HD的小鼠模型，并更好地了解细胞重编程在淋巴肿大中的作用。第二种EBV相关疾病是移植后淋巴增殖性疾病(PTLD)，在移植后患者中很常见，很大程度上是由于EBV感染的B细胞由于免疫抑制而产生的。我们发现，在小鼠发育中的B细胞中，EBV蛋白LMP1的诱导表达导致免疫系统对这些细胞的排斥反应。LMP1是EBV介导的B细胞转化的主要参与者。动物体内T细胞的耗尽导致表达LMP1的B细胞迅速生长，小鼠在几周内死亡。我们计划将该系统开发成第一个PTLD小鼠模型，通过将LMP1表达靶向成熟的B细胞，并充分描述突变动物的疾病进展和可逆性，以期最终利用该模型开发新的治疗策略。公共卫生相关性：霍奇金氏病(HD)是西方世界最常见的淋巴瘤，而移植后淋巴增生性疾病(PTLD)是移植后患者发病率和死亡率的重要原因，是一个重要的临床问题。EB病毒(Epstein-Barr-Virus，EBV)在HD和PTLD中起主要作用，近一半的HD和PTLD与EBV相关。目前的建议是基于这些疾病在人类中的发病机制的已知特征和广泛的自己的工作，并旨在建立与EBV相关的HD和PTLD小鼠模型，这两种模型迄今尚不存在，但应为新的治疗策略开辟道路，并为基因表达重新编程在淋巴肿瘤发生中的作用提供新的线索。

DESCRIPTION (provided by applicant): In Hodgkin's Disease (HD), the most common lymphoma in the Western world, the malignant cells are the so-called Hodgkin and Reed-Sternberg (HRS) cells, comprising only a few percent of the lymphoma mass. HRS cells are often infected by Epstein-Barr-Virus (EBV) and express the EBV proteins LMP1 and LMP2A, partially mimicking constitutively an active CD40 co-receptor and B cell antigen receptor (BCR), respectively. Previous work, which had identified somatically mutated B cells, often bearing mutations "crippling" BCR expression, as HRS progenitors, led to a scenario of HD pathogenesis, in which HRS cells derive from pre-apoptotic GC B cells rescued by LMP2A and LMP1 expression. Despite their B cell origin, HRS cells have largely lost the B cell-specific gene expression program and acquired expression of genes typical for the T and/or myeloid hematopoietic lineages. We hypothesize that this lineage infidelity is dictated by the interference of certain transcription factors (TFs) known to be expressed in HRS cells, namely Id2, ABF1 and Notch1, with the B cell- specific gene expression program. This together with constitutive expression of proteins promoting cell survival and proliferation, like the TF c-Jun and TFs of the NF-kB family, may ultimately reprogram GC B cells into HRS cells. In this general scenario, EBV infection is a major initial transforming event in HD, and is indeed known to result in up-regulation of the TFs NF-kB, AP1, Id2 and ABF1 in B cells. Following this general hypothesis, we have developed a strategy to target the expression of candidate genes into GC B cells, using Cre-mediated conditional gene targeting. Using and further improving these tools, we plan to analyze whether induced expression of the various EBV-derived proteins and TFs individually and in combination in GC B cells will result in trans-differentiation and transformation of those cells as it is seen in HRS cells in HD. Our ultimate goal is to generate a mouse model of HD and to better understand the role of cellular reprogramming in lymphomagenesis. A second EBV-associated disease addressed in the present proposal is Post-Transplant Lymphoproliferative Disorder (PTLD), common in post-transplantation patients and due to a significant extent to the outgrowth of EBV-infected B cells because of immune suppression. We have found that induced expression of the EBV protein LMP1, known to be a major player in EBV-mediated B cell transformation, in developing B cells in the mouse leads to the rejection of these cells by the immune system. Depletion of T cells in the animals results in the rapid outgrowth of LMP1 expressing B cell blasts and death of the mice within a few weeks. We plan to develop this system into a first mouse model of PTLD, by targeting LMP1 expression into mature B cells, and to fully characterize disease progression and reversibility in the mutant animals, with a view of ultimately using this model for the development of new therapeutic strategies. Public Health Relevance: Hodgkin's Disease (HD) is the most common lymphoma in the Western world, and Post-Transplant Lymphoproliferative Disorder (PTLD) is an important cause of morbidity and mortality in post-transplantation patients, posing a significant clinical problem. Epstein-Barr-Virus (EBV) plays a major role in both HD and PTLD, with almost half of the HD cases and most of PTLDs being associated with it. The present proposal is based on known features of the pathogenesis of these diseases in the human and extensive own work, and aims at the generation of EBV-related mouse models of HD and PTLD, which so far do not exist, but should open the way to new therapeutic strategies and also shed new light on the role of gene expression reprogramming in lymphomagenesis.