CLONAL EVOLUTION OF STRUCTURAL VARIANTS IN THE PROGRESSION OF METASTATIC COLORECT

描述（由申请人提供）：直到最近，癌症研究界一直在努力发现上皮癌中的基因融合。然而，下一代测序（NGS）的进展表明，可以产生基因融合转录物的结构变异（SVs）是在实体肿瘤中普遍存在的一类严重缺乏代表性的突变。从分析的角度来看，我们面临的关键挑战是确保偶然的基因融合不仅可以被检测到，而且可以在癌症转录组中越来越多的嵌合突变中相应地优先考虑。此外，了解基因融合在肿瘤进展和治疗反应中的作用将具有直接的临床影响。目前，NGS已经提供了肿瘤内寡克隆组成和进化的高分辨率快照。基因组研究所的开创性研究利用全基因组测序（WGS）发现的单核苷酸变异（snv）进行深度靶向测序，计算变异等位基因频率（VAFs），以研究克隆随时间的进化。然而，现有的克隆进化研究只关注snv，而忽略了SVs及其相应的基因融合产物。无法将基因融合纳入肿瘤细胞的克隆分析是该领域的一个关键障碍。此外，与血液学恶性肿瘤不同，重复活检是常见的，而实体瘤患者连续活检并不常见，从而限制了我们监测患者的能力。为了避免这个问题，最近的努力集中在检测血浆中循环肿瘤DNA （ctDNA）的基因组畸变上。鉴于基因融合在人类癌症中的临床意义，本提案的重点是通过以下目标提高我们在肿瘤进化过程中克隆群体背景下检测和监测基因融合的能力：(1)整合SV-Seq和ChimeraScan，建立一种强大的计算方法，从WGS和RNA-Seq数据中检测高可信度的基因融合；(2)在一组结直肠癌患者中应用我们的集成方法，从相应的基因组断点中识别基因融合并计算其VAFs，以了解它们在转移性疾病进展过程中在（创始或亚）克隆中的作用。(3)评估监测ctDNA中的基因融合（通过它们的基因组断点）是否可以概括肿瘤活检中观察到的情况。在这项研究中，考虑到我们独特的患者队列，原发和转移性肿瘤的WGS和RNA-Seq的可用性，以及手术时和随访时收集的血浆的可用性，我们选择专注于结直肠癌。当我们专注于

DESCRIPTION (provided by applicant): Until recently the cancer research community was struggling to discover gene fusions in epithelial cancers. However, advances in Next Generation Sequencing (NGS) have revealed that structural variants (SVs), which can generate gene fusion transcripts, are a severely under-represented class of mutations that are prevalent across solid tumors. From an analysis perspective, we are faced with the critical challenge of ensuring that a casual gene fusion is not only detected, but can be prioritized accordingly amongst the increasing number of chimeric mutations in a cancer transcriptome. Furthermore, it would be of immediate clinical impact to understand the role of gene fusions during tumor progression and in response to treatment. Currently, NGS has provided a high-resolution snapshot of the oligoclonal composition and evolution within tumors. This can be exemplified by pioneering studies at The Genome Institute leveraging deep, targeted sequencing of single nucleotide variants (SNVs) discovered using whole genome sequencing (WGS) to calculate variant allele frequencies (VAFs) for studying clonal evolution over time. However, existing discoveries understanding clonal evolution focused solely on SNVs thereby missing SVs and their corresponding gene fusion products. The inability to incorporate gene fusions into clonal analysis of tumor cells represents a critical barrier in the field. Furthermore, unlike hematologicl malignancies where it is common to take repeat biopsies, it is not commonplace to serially biopsy patients with solid tumors thereby limiting our ability to monitor patients. To circumvent this issue, recent efforts have focused on detecting genomic aberrations from circulating tumor DNA (ctDNA) within the plasma. Given the clinical significance of gene fusions across human cancers, this proposal focuses on improving our ability to detect and monitor gene fusions within the context of clonal populations throughout tumor evolution through the following aims: (1) integrate SV-Seq and ChimeraScan to establish a robust computational method to detect high-confidence gene fusions from WGS and RNA-Seq data, (2) apply our integrated method across a cohort of colorectal cancer patients to identify gene fusions and calculate their VAFs, from their corresponding genomic breakpoints, to understand their role within (founder- or sub-)clones during the progression to metastatic disease, and (3) assess whether monitoring gene fusions (via their genomic breakpoints) in ctDNA can recapitulate what is observed in tumor biopsies. In this study we have chosen to focus on colorectal cancer given our unique patient cohort, the availability of WGS and RNA-Seq from primary and metastatic tumors, and the availability of plasma collected at the time of surgery and from a follow-up visit. While our focus on colorectal cancer may yield novel discoveries, this work is broadly applicable across human cancers. Just as understanding the role of SNVs in clonal evolution has led to groundbreaking discoveries, we envision that this proposal will lay the foundation for similar discoveries focusin on gene fusions that could impact patient care.