UNDERSTANDING THE ROLE OF EPSTEIN BARR VIRUS IN T CELL AND NATURAL KILLER CELL LYMPHOPROLIFERATIONS AND MALIGNANCIES

爱泼斯坦-巴尔病毒(EBV)是一种常见的疱疹病毒，它会在每个人的一生中感染大多数人。绝大多数人没有意识到他们已经感染了病毒，因为他们的免疫系统严格控制着病毒。虽然EBV最为人所知的是引起腺热，但它也与少数人的癌症有关。这些癌症通常发生在作为病毒自然生命周期一部分被感染的细胞、B淋巴细胞和上皮细胞。然而，在极少数情况下，EBV也可以感染与病毒生命周期无关的细胞、T淋巴细胞和自然杀伤(NK)细胞。这些细胞类型的感染总是会导致严重的、危及生命的疾病，包括癌症。这些疾病极难治疗，临床结果非常差，通常以几周到几个月的存活率来衡量。迫切需要更好地了解这些EB病毒相关性T/NK细胞疾病的病理机制，以便开发新的有效的、个性化的治疗方法。由于这些疾病的罕见以及我们无法在实验室感染T/NK细胞，对这些疾病的了解进展极其缓慢。然而，我们已经解决了这两个问题。我们目前有一批来自英国EBV相关T/NK细胞疾病患者的血液样本准备进行分析。此外，我们优化了培养条件，并从患者样本中产生了匹配的EBV阳性T/NK细胞株。最后，也许也是最重要的，我们已经成功地在我们的实验室中复制感染T细胞，这将使我们第一次能够检查EBV是如何触发这些T/NK细胞疾病并诱导它们转化为恶性肿瘤的。我们有四个主要目标。(1)利用患者的血液，我们已经从微环境中确定了帮助患者感染EBV的T/NK细胞在培养中生长所需的白细胞和化学信号(细胞因子)。我们计划在实验室中复制这些条件，以帮助新感染的T/NK细胞生长，并确定EBV、周围细胞和细胞因子如何促进新感染的T/NK细胞的生长。(2)一旦生长条件优化，我们将检测特定的EBV基因产物如何能够刺激EBV相关T/NK细胞疾病中观察到的慢性炎症。(3)通过在我们的细胞培养中复制EBV感染和慢性炎症，我们将研究EBV和慢性炎症是如何驱动感染的细胞无限生长和失调抗EBV免疫的。(4)我们建立了一种新的分析工具，用EBV标志物和细胞类型标志物标记EBV感染的细胞。我们可以扩大标记物的范围，以确定EBV感染细胞上的潜在新药靶点，以包括治疗性抗体，如抗CCR4(Mogamulizumab)，目前正处于外周T细胞淋巴瘤的第二阶段临床试验。此外，如上所述，我们可以在药物存在的情况下培养匹配的体外EBV感染的T/NK细胞，以确定对药物的敏感性。这是第一次通过结合体外患者样本和原发感染的T/NK细胞来分析EBV在EBV相关T/NK细胞疾病中的作用的研究。我们预计，通过了解EBV如何直接或与周围微环境的其他细胞和细胞因子结合导致疾病，并通过监测疾病在体外和体内的进展，我们将更好地理解替代的新治疗方案。最后，根据这项研究的结果，如果这项研究进行试验，我们为检查潜在的新治疗方法而建立的检测方法将对EBV相关T/NK细胞疾病患者群体有更广泛的好处。结果将确定个性化治疗选择的潜在药物。

Epstein Barr virus (EBV) is a common herpesvirus that infects the majority of the human population for the lifetime of each individual. The vast majority of individuals are unaware that they have been infected because the virus is kept under tight control by their immune system. Although EBV is most well known for causing glandular fever, it is also associated with cancers in a small number of individuals. These cancers usually occur in the cells infected as part of the natural life cycle of the virus, B lymphocytes and epithelial cells. However, on rare occasions EBV can also infect cells that are not involved in the lifecycle of the virus, T lymphocytes and Natural killer (NK) cells. Infection of these cell types always results in serious, life-threatening diseases including cancers. These diseases are extremely difficult to treat and have a very poor clinical outcome, usually measured in survival from weeks to months. There is an urgent need to better understand the pathology of these EBV-associated T/NK cell diseases in order to develop new effective, personalized therapies.Progress into understanding these diseases has been extremely slow, due to the rarity of the diseases and our inability to infect T/NK cells in the lab. However, we have addressed both these issues. We currently have a bank of blood samples from UK patients with EBV-associated T/NK cell diseases ready for analysis. Furthermore we have optimized the culture conditions and have generated matching EBV-positive T/NK cell lines from the patients' samples. Finally, and perhaps most importantly, we have managed to reproducibly infect T cells in our laboratory, which for the first time will allow us to examine how EBV triggers these T/NK cell diseases and induces their transformation into malignancy. We have four key objectives. (1) Using patient blood, we have identified white blood cells and chemical signals (cytokines) from the microenvironment which are required to help the patients' EBV-infected T/NK cells to grow in culture. We plan to replicate these conditions in the lab to help the newly-infected T/NK cells grow and determine how EBV, the surrounding cells and cytokines contribute to the growth of newly-infected T/NK cells. (2) Once the growth conditions are optimized, we will examine how specific EBV gene products able to stimulate the chronic inflammation observed in the EBV-associated T/NK cell diseases. (3) By replicating the EBV infection and chronic inflammation in our cell cultures, we will examine how EBV and chronic inflammation drive the infected cells to grow indefinitely and dysregulate the anti-EBV immunity. (4) We have established a new analysis tool, which labels the EBV-infected cells with markers for EBV and markers for the cell type. We can expand the range of markers to identify potential new drug targets on the EBV-infected cell to include therapeutic antibodies such as anti-CCR4 (mogamulizumab) currently in phase 2 clinical trial for peripheral T cell lymphoma. Furthermore, we can culture the matching ex-vivo EBV-infected T/NK cells, as above, in the presence of the drug to determine sensitivity to the drug. This is the first study of its kind to analyse the contribution of EBV to the EBV-associated T/NK cell diseases by combining ex-vivo patient samples and primary infected T/NK cells. We anticipate that by understanding how EBV causes the diseases, either directly or in combination with other cells and cytokines of the surrounding microenvironment, and by monitoring disease progression both in vitro and in vivo, we will develop a better understanding of alternative novel therapeutic options. Finally, the assays we have established to examine the potential new treatments will have wider benefit to the EBV-associated T/NK cell disease patient groups if the study were taken to trial, based on the findings of this study. The results will identify potential drugs for personalized treatment options.