IMAT-ITCR Collaboration: Development of a high-resolution mapping platform for HPV DNA integration in premalignant lesions

摘要/摘要 病毒在10%到20%的人类癌症中扮演着重要角色。将特定病毒与人类癌症联系在一起仍然是一个 基本的生物医学和技术问题。一个病毒家族，人乳头瘤病毒(HPV)， 包括225多种已知类型和引起的病变，从良性疣到高度致命的侵袭性病变 癌症。HPV诱导的肿瘤发生在几个解剖部位，包括几乎100%的宫颈 癌症，90%的肛门癌，30%到60%的头颈部肿瘤，以及大约四分之一到一半的阴道肿瘤， 外阴癌和阴道癌。在大多数HPV诱导的肿瘤中，至少部分病毒DNA基因组已经变成 整合到人类基因组中，可能是宿主细胞DNA修复过程异常的结果。 我们的实验室工作一直致力于开发准确、灵敏、广谱的特异性技术 基于DNA杂交捕获和大规模并行测序的检测和结构分析 癌前病变和浸润性肿瘤中HPV DNA的整合。正在进行的研究也在不断发展 荧光显微镜技术同时检测整合的HPV DNA和HPV RNA转录本，AS 这些为检测HPV诱导的肿瘤细胞的存在提供了高度特异性和潜在的诊断工具， 包括治疗后残留疾病的临床样本。我们的工作包括远程纳米孔 DNA测序和RNA测序以确认HPV和 人类基因组和阐明病毒对病毒和人类基因转录的潜在影响。这些问题的关键 HAAS小组(麻省理工学院的博德研究所)与 Lenz(阿尔伯特·爱因斯坦医学院)和Montanna(新泽西州罗格斯癌症研究所)实验室 扩展三位一体癌症转录组分析工具包(CTAT)的计算工具。这导致了 用于检测整合HPV DNA、结构组装的扩展CTAT组件的研制 整合的病毒DNA片段，以及HPV诱导的肿瘤和 癌前病变。GitHub上提供了CTAT病毒插入查找器(CTAT-VIF)。IMAT-ITCR 此处建议的协作将通过实现另外三个目标来显著扩展正在进行的协作 目标。AIM 1将扩展CTAT-VIF以分析约13,000种不同病毒的存在并调查是否 任何病毒DNA都是通过筛选所有癌症基因组图谱(TCGA)和 基因-组织表达项目(GTEx)数据集。目标2是鉴定和分析新型病毒--人类 病毒DNA插入产生的融合转录本。AIM 3将扩大我们对HPV的现场分析- 诱导宫颈癌和癌前病变包括空间转录分析，包括 为这项技术扩展CTAT。这些研究应该改进计算和实验室 了解病毒在人类癌症中的作用的技术。

Abstract/Summary Viruses play a role in 10 to 20% of human cancers. Tying specific viruses to human cancers remains a fundamental biomedical and technological problem. One virus family, the human papillomaviruses (HPVs), comprises over 225 known types and causes lesions ranging from benign warts to highly lethal, invasive carcinomas. HPV-induced tumors occur at several anatomical sites including nearly 100% of cervical cancers, 90% of anal cancers, 30 to 60% of head & neck tumors, and roughly one-fourth to one-half of vaginal, vulvar & penile cancers. In most HPV-induced tumors, at least part of the viral DNA genome has become integrated into the human genome, presumably as a consequence of aberrant host cell DNA repair processes. Our laboratory work has been focused on the development of accurate, sensitive, broad specificity technology based on DNA hybridization capture plus massively parallel sequencing to detect and structurally analyze integrated HPV DNA in precancerous lesions and invasive tumors. Ongoing studies are also developing fluorescent microscopy technologies for detection of both integrated HPV DNA and HPV RNA transcripts, as these provide highly specific, potential diagnostic tools to detect the presence of HPV-induced tumor cells, including post-therapy residual disease in clinical samples. Our work encompasses long-range, nanopore DNA sequencing plus RNA sequencing to confirm any complex structural rearrangements of the HPV and human genomes and to elucidate potential viral effects on viral and human gene transcription. Key to these studies has been a very successful, collaborative effort by the Haas group (Broad Institute at MIT) with the Lenz (Albert Einstein College of Medicine) and Montagna (Rutgers Cancer Institute of New Jersey) labs to expand the computational tools of the Trinity Cancer Transcriptome Analysis Toolkit (CTAT). This has led to the development of expanded CTAT components for detection of integrated HPV DNA, structural assembly of integrated viral DNA segments, and overall transcriptional analysis of HPV-induced tumors and precancerous lesions. The CTAT-virus insertion finder (CTAT-VIF) is available on GitHub. The IMAT-ITCR collaboration proposed here will substantially expand the ongoing collaboration by pursuing three additional aims. Aim 1 will expand CTAT-VIF to analyze the presence of ~13,000 different viruses and investigate if any viral DNAs are integrated and/or expressed by screening all The Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression Project (GTEx) datasets. Aim 2 is to identify and analyze novel virus-human fusion transcripts generated from virus DNA insertions. Aim 3 will broaden our in-situ analysis of HPV- induced cervical cancers and precancerous lesions to include spatial transcriptomic analysis, including expanding CTAT for this technology. These studies should improve computational and laboratory technologies for understanding the roles of viruses in human cancer.