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）的进展已经揭示，可以产生基因融合转录物的结构变体（SV）是在实体瘤中普遍存在的严重代表性不足的一类突变。从分析的角度来看，我们面临着确保偶然基因融合不仅被检测到，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的关键挑战。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。这可以通过基因组研究所的开创性研究来举例说明，该研究利用全基因组测序（WGS）发现的单核苷酸变异（SNV）的深度靶向测序来计算变异等位基因频率（VAF），以研究随时间的克隆进化。然而，了解克隆进化的现有发现仅关注SNV，从而错过了SV及其相应的基因融合产物。不能将基因融合纳入肿瘤细胞的克隆分析是该领域的关键障碍。此外，不像血液恶性肿瘤，其中进行重复活检是常见的，连续活检患有实体瘤的患者并不常见，从而限制了我们监测患者的能力。为了规避这个问题，最近的努力集中在检测血浆内循环肿瘤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，该提案的重点是通过以下目标提高我们在整个肿瘤进化过程中的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以建立稳健的计算方法来从WGS和RNA-Seq数据检测高置信度基因融合体，（2）将我们的综合方法应用于结直肠癌患者队列，以识别基因融合并从其相应的基因组断点计算其VAF，以了解其在结直肠癌中的作用。（建立者-或亚-）克隆在转移性疾病的进展过程中，和（3）评估ctDNA中的基因融合（通过它们的基因组断裂点）监测是否可以概括在肿瘤活检中观察到的。在这项研究中，我们选择专注于结直肠癌，因为我们独特的患者队列，原发性和转移性肿瘤的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。尽管我们的重点 研究结直肠癌可能会产生新的发现，这项工作广泛适用于人类癌症。正如了解SNV在克隆进化中的作用导致了突破性的发现一样，我们设想这项提议将为类似的发现奠定基础，这些发现集中在可能影响患者护理的基因融合上。

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.