Comprehensive genomic profiling of Hodgkin Lymphoma employing liquid biopsies

霍奇金淋巴瘤是一种B细胞来源的血液系统恶性肿瘤。发病高峰在成年早期。霍奇金淋巴瘤的显著特征之一是，病理性恶性霍奇金和Reed-Sternberg细胞仅占肿瘤组织的一小部分（1-10%），这对全面的基因组分析研究提出了重大的技术挑战。首先，所需的积极、多药剂、一线治疗会导致许多早期和晚期毒性，例如继发性癌症、心血管疾病、不孕症、疲劳和骨坏死。第二，10-30%的患者在第一次治疗尝试中失败，这些通常非常年轻的患者中约有一半最终死亡。最近，我们建立了一个基于液体活检的靶向测序策略，用于霍奇金淋巴瘤的微小残留病诊断。我们发现，考虑到活检中肿瘤细胞的缺乏，霍奇金淋巴瘤中的无细胞血浆DNA高度富集肿瘤来源的DNA。因此，我们使用定制的全外显子组测序管道进行了一项试点研究，以评估使用液体活检进行霍奇金淋巴瘤综合基因组分析研究的可能性。我们能够证明这是一种可行的方法，可以检测新的体细胞改变，并全面了解霍奇金淋巴瘤的基因组学。我们现在建议将我们的方法扩展到150例临床试验患者，这些患者已经获得了材料和广泛的临床数据，包括结果。我们的目标是使用一种新的内部开发的液体活检测序平台来创建新诊断的霍奇金淋巴瘤的第一个大的、完全临床注释的综合基因组谱，并将其用于（I）发现霍奇金淋巴瘤中的新的遗传改变，因为在霍奇金淋巴瘤中仍然存在对综合基因组谱研究的未满足的需求，（II）将复发性生殖系和体细胞变体与全面可用的临床数据相关联，（III）根据复发性变异，开发新的霍奇金淋巴瘤风险分类（例如，根据遗传信息识别高风险或低风险患者，以便在未来进行个性化和优化治疗），（IV）通过捕获所有患者肿瘤基因组的快照来了解患者疾病的克隆组成和异质性，以及（V）证明了液体活检的全外显子组测序用于临床使用和治疗个体化的可行性，其相关性超出了霍奇金淋巴瘤本身。我们将通过可用肿瘤活检的全外显子组测序和使用所提出的全外显子组测序方法和我们已经开发的样本子集的独立双重分析来全面验证我们的发现，也可以在非常高的覆盖率（> 1500 x）下对霍奇金淋巴瘤的选定复发性遗传改变进行基于液体活检的靶向测序策略。

Hodgkin lymphoma is a B cell-derived, haematological malignancy. The peak incidence is in early adulthood. One of the remarkable features of Hodgkin lymphoma is the fact that the pathognomonic, malignant Hodgkin and Reed-Sternberg cells make up only a small percentage (1-10%) of the tumour tissue which poses a major technical challenge to comprehensive genomic profiling studies.Two challenges define the current medical need in Hodgkin lymphoma. First, the needed aggressive, multi-agent, front-line treatment leads to many early and late toxicities such as secondary cancers, cardiovascular disease, infertility, fatigue and osteonecrosis. Second, 10-30% of patients fail the first treatment attempt and approximately half of these often very young patients will ultimately die. Recently, we have established a liquid biopsy based targeted sequencing strategy for minimal residual disease diagnostics in Hodgkin lymphoma. We found that cell-free plasma DNA is highly enriched for tumour-derived DNA in Hodgkin lymphoma, considering the paucity of tumour cells in biopsies. We therefore performed a pilot study using a customized whole exome sequencing pipeline to evaluate the possibility of a comprehensive genomic profiling study of Hodgkin lymphoma using liquid biopsies. We were able to show that this is a feasible approach that can detect novel somatic alterations and comprehensively inform on the genomics of Hodgkin lymphoma.We now propose to extend our approach to 150 clinical trial patients for whom material and extensive clinical data including outcome is already available. We aim to use a novel, in-house developed liquid biopsy sequencing platform to create the first large, fully clinically annotated comprehensive genomic profile of newly diagnosed Hodgkin lymphoma and use it to (I) find novel genetic alterations in Hodgkin lymphoma as there is still an unsatisfied demand of a comprehensive genomic profiling study in Hodgkin lymphoma, (II) correlate both recurrent germline and somatic variants with comprehensively available clinical data, (III) develop novel risk classifications of Hodgkin lymphoma based on recurrent variants (e.g. identify high-risk or low-risk patients based on genetic information allowing to individualize and optimize treatment in the future), (IV) understand the clonal composition and heterogeneity of a patient's disease by capturing a snapshot of all the patient's tumour genome and (V) prove the feasibility of whole exome sequencing of liquid biopsies for clinical use and treatment individualization with relevance beyond Hodgkin lymphoma itself.We will comprehensively validate our findings with whole exome sequencing of available tumour biopsies and an independent dual analysis of a subset of samples with both the proposed whole exome sequencing approach and our already developed, also liquid biopsy based targeted sequencing strategy of selected recurrent genetic alterations of Hodgkin lymphoma at very high coverage ( > 1500x).