Identifying recurrent driver mutations in skin cancers by targeted UV damage sequencing

通过靶向紫外线损伤测序识别皮肤癌中的复发性驱动突变

• 批准号: 10645759
• 负责人: John J Wyrick
• 金额: $ 25.66万
• 依托单位: WASHINGTON STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-10 至 2025-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10645759
• 关键词: Adenine; Affect; BRAF gene; Binding; Binding Sites; Bioinformatics; Cancer Etiology; Cancer cell line; Cell Line; Cell Proliferation; Cell Survival; Cells; Codon Nucleotides; DNA; DNA Binding; DNA Damage; DNA mapping; Data; Data Set; Early Promoters; Elements; Excision Repair; Exons; Exposure to; Family; Frequencies; Gene Proteins; Genes; Genome; Growth; Human; Human Genome; In Vitro; Individual; Lesion; Link; Malignant Neoplasms; Malignant neoplasm of urinary bladder; Maps; Measures; Methods; Molecular; Mutate; Mutation; Nucleotide Excision Repair; Nucleotides; Oncogenic; Pathway interactions; Patients; Phosphotransferases; Proteins; Protocols documentation; Publishing; Pyrimidine; Pyrimidine Dimers; Recurrence; Resolution; Role; Sampling; Site; Skin; Skin Cancer; Testing; Thymine; Triage; Tumor-Derived; UV Radiation Exposure; UV induced; Ultraviolet Rays; Untranslated RNA; Yeasts; base; bioinformatics pipeline; cancer cell; cancer genome; cancer type; candidate identification; carcinogenesis; carcinogenicity; driver mutation; endonuclease; genome sequencing; genomic locus; keratinocyte; melanocyte; melanoma; melanomagenesis; repaired; response; transcription factor; tumor; ultraviolet; ultraviolet damage; ultraviolet irradiation

Abstract: Skin cancers, such as melanoma, are among the most mutated human cancers due to the mutagenic action of ultraviolet (UV) light. The high mutation load in skin cancers makes it challenging to identify driver mutations in these tumors, particularly in non-coding DNA. For example, recent genome sequencing efforts have identified dozens of recurrent exon mutations and hundreds of recurrent non-coding mutations in melanomas. However, current evidence suggests that only a subset of these are under carcinogenic selection and contribute to melanomagenesis. Many recurrent mutation sites are found in the DNA-binding sites of E26 transformation- specific (ETS) family of transcription factors (TFs). We and others have recently shown that ETS and other TFs induce high levels of UV damage, both in UV-exposed cells and in vitro, which can potentially explain the presence of recurrent mutations at their binding sites. To test this hypothesis, we have developed new capture- sequencing methods for mapping UV damage at targeted sites in the human genome. We propose to use these methods to map DNA damage in UV-irradiated skin cells at sites of recurrent mutations in skin cancers. The overall objective of this proposal is to use targeted UV damage sequencing to distinguish between recurrent mutations that are simply caused by elevated levels of UV damage (often at TF binding sites) from those that cannot be explained by UV damage, and therefore are more likely to be oncogenic mutations. In Aim I, we will use the CPD-capture-seq method to map the formation and repair of UV-induced cyclobutane pyrimidine dimers (CPDs) in UV-irradiated primary skin cells and cancer cell lines. In parallel, will develop a new method, known as UVDE-capture-seq, to map less common 6-4 photoproducts (6-4PPs) and atypical photoproducts at sites of recurrent mutations in UV irradiated cells. In Aim II, we will develop a bioinformatics pipeline to use these capture sequencing data sets to identify candidate non-coding driver mutations by triaging recurrent mutations associated with elevated UV damage. While it is possible that a few of the recurrent mutations associated with elevated UV damage could contribute to carcinogenesis, multiple lines of evidence suggest that the vast majority of these are passenger mutations. In parallel, we will functionally characterize candidate non-coding driver mutations identified from our preliminary CPD-capture-seq data. We will also determine whether UV damage induction due to transient ETS TF binding causes a subset of recurrent exon mutations in melanoma and other skin cancers. Successful completion of these aims will provide a new method for identifying driver mutations in skin cancers, particularly in non-coding DNA. Importantly, the methods developed in this proposal could be adapted to analyze different classes of DNA damage associated with recurrent mutation sites in other cancer types.

抽象的： 皮肤癌，例如黑色素瘤，是由于人类最突变的癌症之一 紫外线（UV）光。皮肤癌的高突变负荷使识别驾驶员突变的挑战 这些肿瘤，特别是在非编码DNA中。例如，最近的基因组测序工作已经确定 黑色素瘤中的数十个复发外显子突变和数百种复发的非编码突变。然而， 当前的证据表明，其中的一部分仅在致癌中，并有助于 黑素作仿。在E26转化的DNA结合位点发现了许多复发位点 - 特定（ETS）转录因子家族（TFS）。我们和其他人最近表明了ET和其他TF 在紫外线暴露的细胞和体外诱导高水平的紫外线损伤，这可能会解释 在其结合位点存在复发突变。为了检验这一假设，我们开发了新的捕获 - 在人基因组中靶向位点映射紫外线损伤的测序方法。我们建议使用 这些方法是在皮肤癌中复发突变部位的紫外线辐照皮肤细胞中绘制DNA损伤的方法。 该提案的总体目的是使用有针对性的紫外线损伤测序来区分 仅由紫外线损伤水平升高（通常在TF结合位点）引起的复发突变 那些不能用紫外线损伤来解释的，因此更可能是致癌突变。在 AIM I，我们将使用CPD捕获式方法来绘制紫外线诱导的环丁烷的形成和修复 在紫外线辐照的原代皮细胞和癌细胞系中，嘧啶二聚体（CPD）。并行，会发展 新方法，称为UVDE捕获式seq，以绘制较少常见的6-4个光产物（6-4pps）和非典型的绘制 紫外线辐照细胞复发突变部位的光产物。在AIM II中，我们将开发生物信息学 使用这些捕获测序数据集的管道来识别候选非编码驱动程序突变 与紫外线损伤升高有关的复发突变。虽然有可能 与紫外线损伤升高有关的复发突变可能导致癌变，多条线 证据表明，其中绝大多数是乘客突变。同时，我们将在功能上 表征从我们的初步CPD捕获率数据确定的候选非编码驱动器突变。我们 还将确定由于瞬态ETS TF结合引起的紫外线损伤是否会导致子集 黑色素瘤和其他皮肤癌中的复发外显子突变。这些目标的成功完成将 提供了一种鉴定皮肤癌中驱动突变的新方法，尤其是在非编码DNA中。 重要的是，可以对本提案中开发的方法进行调整以分析不同类别的DNA 与其他癌症类型的复发突变部位相关的损害。