Pre-BCR Function Selecting Novel B Cell Receptors in Chronic Lymphocytic Leukemia

描述（由申请人提供）：本申请涉及广泛的挑战领域（06）使能技术和特定的挑战主题，06- ca -117：癌症发展，病理学和病理进展。B细胞慢性淋巴细胞白血病（CLL）是西方国家最常见的成人白血病，它是由B系细胞的克隆群体积累而成的，其表面表型与大多数成熟B细胞不同。虽然这种疾病已经被发现30多年了，但令人惊讶的是，其病因尚不清楚，20年前建立的标准治疗方法会产生明显的副作用，而且不会影响治愈，反而会延迟进展。在过去的十年中，根据B细胞抗原受体（BCR）的突变程度，CLL可以细分为两类：显著突变（> %）的CLL表现出较慢的进展，通常是良性的过程，而很少突变的CLL预后更差，进展更快。对BCR重链可变区（VHs）的序列分析揭示了有趣的发现，表明VH基因的使用存在显著偏差，并且相似甚至相同的CDR3片段的重复使用，这表明在cll或其前体的发展中，共同抗原的选择发挥了作用。也有明确的证据表明，CLL bcr通常编码自身反应性，这种自身反应性可能在疾病过程中发挥关键作用。此外，最近的研究描述了CLL患者亲属中CLL样单克隆扩增的存在，但也更普遍地存在于60岁以上的成年人中。由于CLL是一种发病率随年龄增长而增加的疾病，很少在年轻人中发现，因此我们很容易推测克隆B细胞通常随着年龄的增长而积累，但很少进展到CLL阶段。了解产生这些单克隆b细胞淋巴细胞（MBLs）的因素并确定它们如何发展为CLL可能会导致CLL治疗的突破。在过去的二十年里，我们的兴趣一直集中在开发和表征一种小鼠自身反应性b细胞，这种细胞可能会成为MBL和最终CLL的倾向。这些细胞最初通过表达CD5（以前在小鼠中称为Ly-1）来鉴定，显示出独特的定位和功能，重要的是在小鼠中负责许多天然自身抗体的产生。若干年后，我们（和其他人）发现在人CD5+ b细胞中，自身抗体类风湿性因子的产生是丰富的。我们现在认为，时机已经成熟，可以探讨小鼠和人类CD5+ B细胞和CD5+ B细胞白血病中bcr的V基因中出现的显著偏差，是否至少在一定程度上涉及一种尚未被认识的机制，这种机制塑造了自然自身反应性B细胞的全部功能。也就是说，在pre-B发育阶段，存在基于pre-BCR组装的强选择，并且这种_selection检查点可能对理解具有自反应性偏倚的独特重链的发育很重要。我们已经开发了使用小鼠细胞来测量Ig重链组装成预bcr的能力的实验，并将其与诱导增殖相关联。我们现在建议扩展这项工作，检查在一系列晚期发展的cll样小鼠白血病中使用的重链，但重要的是也建立与人类细胞可比较的测定方法。这将使我们能够评估人体系统中的前bcr组装和信号传导，然后检查CLL和MBL的重链，以询问在这些细胞中观察到的独特的重链偏差是否在发育早期出现，可能为这类细胞的生长和最终转化提供重要的新见解。我们提出的工作将为进一步研究BCR在CLL前体发展中的作用奠定基础，最终发现新的治疗方法。最后，这个项目的一个重要方面是，它将运用我们在小鼠B细胞发育、选择和亚群方面的丰富经验和专业知识，集中研究人类白血病的潜在病因，具有开发更好、更有针对性的治疗方法的潜力。

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (06) Enabling Technologies, and specific Challenge Topic, 06-CA-117: Cancer Development, Pathology, and Pathological Progression. B cell chronic lymphocytic leukemia (CLL) is the most common leukemia affecting adults in Western countries and is recognized by the accumulation of a clonal population of B lineage cells with a surface phenotype distinguishing them from most mature B cells. Although the disorder has been know for over thirty years, surprisingly, its etiology remains unclear and standard treatments, established twenty years ago, entail significant side effects and do not affect a cure, but rather delay progression for a time. In the past ten years it has become clear that CLL can be subdivided into two categories based on extent of mutation of the B cell antigen receptor (BCR): those with significant mutation (>2%) show slower progression and generally a more benign course, whereas those with little mutation have a much worse prognosis and faster progression. Sequence analysis of heavy chain variable regions (VHs) of the BCR has revealed intriguing findings indicating significant VH gene usage biases, and the recurrent usage of similar or even identical CDR3 segments, suggesting a role for selection by common antigens in development of either CLLs or their precursors. There is also clear evidence that CLL BCRs often encode autoreactivity and such self-reactivity may play a key role in the course of disease. Furthermore, recent studies have described the presence of CLL-like monoclonal expansions in relatives of CLL patients, but also more generally in adults over the age of 60. As CLL is a disease whose incidence increase with age, rarely found in young individuals, it is tempting to speculate that clonal B cells often accumulate with age, but only infrequently progress to the CLL stage. Understanding the factors that generate these monoclonal B-cell lymphocytoses (MBLs) and determining how they progress to CLL could lead to breakthroughs in the treatment of CLL. Our interest over the past twenty years has focused on development and characterization of one kind of autoreactive B-cell in the mouse that may model the cell type with a propensity to become MBL and eventually, CLL. These cells, initially identified by expression of CD5 (previously called Ly-1 in mouse), show distinctive localization and functions, importantly being responsible for much of the natural autoantibody production in mice. Some years later, we (and others) showed that production of autoantibody rheumatoid factor was enriched in human CD5+ B-cells. We now think the time is ripe to ask whether generation of the striking biases seen in V genes of BCRs from both mouse and human CD5+ B-cells and CD5+ B-cell leukemias may involve, at least in part, an as-yet un-appreciated mechanism that sculpts the repertoire of natural autoreactive B cells. That is, at the pre-B stage of development, there is strong selection based on assembly of a pre-BCR, and this _-selection checkpoint may be important in understanding the development of distinctive heavy chains with an autoreactive bias. We have developed assays using mouse cells for measuring the capacity of Ig heavy chains to assemble into a pre-BCR and correlated this with the induction of proliferation. We now propose to extend this work, examining heavy chains used in a series of late-developing CLL-like mouse leukemias, but importantly also establishing comparable assays with human cells. This will allow us to assess of pre-BCR assembly and signaling in the human system, and then examine heavy chains from CLL and MBL, to ask whether the distinctive heavy chain biases seen in these cells arise much earlier in development, potentially providing important new insights into the growth and eventual transformation of such cells. Our proposed work will establish the groundwork needed for further study of the role of BCR in development of CLL precursors, eventually uncovering new treatments. Finally, an important aspect of this project is that it will apply our extensive experience and expertise in mouse B cell development, selection, and subsets in a focused study of a potential etiology of a human leukemia, with the potential of developing better, more targeted, therapies. PUBLIC HEALTH RELEVANCE: This project seeks to understand whether chronic lymophocytic leukemias (CLL), the most common leukemia in adults, arises from a distinct set of natural autoreactive B cells, by testing a property of the antibody present on such cells that distinguishes them from other types of B cells. This will be done using a mouse model for CLL, and also by obtaining patient leukemic cells and directly testing their antibodies. The knowledge generated in this study should provide insights into the origins of CLL and lead to the development of new, more targeted, therapies, improving patient quality of life and potentially even providing a cure for this disease.