TOWARDS A MOUSE MODEL OF CLASSICAL HODGKIN'S DISEASE

描述(申请人提供)：经典型霍奇金氏病(HD)是西方世界最常见的淋巴瘤。HD中的恶性细胞是所谓的霍奇金和里德-斯特恩伯格(HRS)细胞，它们只占淋巴瘤组织的百分之几或更少。在大约一半的患者中，HRS细胞感染了EB病毒(EBV)，并表达EBV编码的膜蛋白LMP1和LMP2A。这些蛋白在结构上具有活性，在B细胞中分别部分模拟CD40共受体和抗原受体(BCR)的信号。基于这些情况和我们自己对显微操作HRS细胞中Ig基因重排的分子分析，我们提出了HD的发病机制。在这种情况下，HRS细胞在大多数情况下来自于某些转化事件挽救的生发中心(GC)前B细胞(S)。在EBV+HD中，LMP1和LMP2a可能参与了最初的救援。这项提议的目的是通过有条件的基因打靶技术在小鼠身上重建这种场景。我们已经开发出一种小鼠突变体，其中Cre重组酶在GC细胞中有效表达，但不能在幼稚B细胞中表达。这将被用来在体内靶向表达LMP2A和/或LMP1到GC B细胞。将调查病毒蛋白对GC反应的干扰。鉴于LMP1的已知致癌特性，可能会观察到由于体细胞过度突变而失去bcr表达的凋亡前GC B细胞的挽救，以及淋巴肿大。作为这些实验的补充，将尝试将HRS细胞的另一个潜在的肿瘤决定因素靶向GC B细胞，即激活形式的Notch1。这种分子最近被证明在HRS细胞中以激活的形式高水平表达。NOTCH1在这方面特别有吸引力，因为它参与淋巴细胞前体细胞的谱系决定，促进T细胞发育。奇怪的是，HRS细胞下调了许多B细胞特异性基因的表达，并表达了包括T细胞在内的其他造血系统的分子标记。NOTCH1，如果异位表达，也是一个有效的癌基因，因此可能有助于这种奇怪的表型以及HRS细胞的转化。通过条件基因打靶将LMP2A、LMP1和Notch1在GC B细胞中的表达结合起来，可能会导致HD小鼠模型的建立。除了淋巴肿大，拟议的实验还应该导致对EBV感染背景下GC反应的生物学的新的见解。

DESCRIPTION (provided by applicant): Classical Hodgkin's disease (HD) is the most common lymphoma in the Western World. The malignant cells in HD are the so-called Hodgkin and Reed-Sternberg (HRS) cells, which comprise only a few percent or less of the lymphoma tissue. In roughly half of the patients, the HRS cells are infected with Epstein-Barr-Virus (EBV) and express the EBV-encoded membrane proteins LMP1 and LMP2A. These proteins are constitutively active and in B cells partially mimic signals of the CD40 co-receptor and the antigen receptor (BCR), respectively. Based on these circumstances and on our own molecular analysis of Ig gene rearrangements in micro manipulated HRS cells we have developed a scenario of HD pathogenesis. In this scenario, HRS cells derive in most instances from pre-apoptotic germinal center (GC) B cells rescued by some transforming event(s). In EBV+ HD, LMP1 and LMP2A may participate in this initial rescue. The aim of this proposal is to reconstruct this scenario in the mouse by conditional gene targeting techniques. We have developed a mouse mutant in which Cre recombinase is efficiently expressed in GC but not naive B cells. This will be used to target expression of LMP2A and/or LMP1 to GC B cells in vivo. The interference of the viral proteins with the GC reaction will be investigated. Rescue of pre-apoptotic GC B cells that have lost BCR expression because of somatic hypermutation might be observed, as well as lymphomagenesis, given the known oncogenic properties of LMP1. These experiments will be complemented by an attempt to target another potential tumor determinant of HRS cells into GC B cells, namely the activated form of Notch1. This molecule has recently been shown to be expressed in HRS cells at high levels and in an activated form. Notch1 is particularly attractive in this context, because it is involved in lineage decisions in lymphocyte progenitors, promoting T cell development. Curiously, HRS cells have down regulated many B cell-specific genes and express molecular markers of other hematopoietic lineages, including T cells. Notch1, which is also a potent oncogene if ectopically expressed, might thus contribute to this curious phenotype as well as to HRS cell transformation. Combining LMP2A, LMP1 and Notch1 expression in GC B cells by conditional gene targeting might lead to a mouse model of HD. Apart from lymphomagenesis, the proposed experiments should also lead to new insights into the biology of the GC reaction in the context of EBV infection.