Transcriptional Signatures of Homologous Recombination Deficiency for Targeted Ch

目标 Ch 同源重组缺陷的转录特征

基本信息

批准号：
8424340
负责人：
Robert W Sobol
金额：
$ 24.35万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-11-15 至 2014-10-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8424340
关键词：
Address Aging BRCA1 gene Base Excision Repairs Biological Markers Bromouridine Cell Line Cell physiology Cells Cisplatin Code DNA Repair DNA Repair Disorder DNA Repair Gene DNA Repair Pathway DNA repair protein DNA-PKcs Defect Development Disease Epigenetic Process Epithelial Epithelial Cells Excision Repair Exposure to Gene Expression Gene Expression Profile Genes Genetic Genetic Transcription Genome Genome Stability Genotoxic Stress Germ Lines Glioma Goals Growth Human Ionizing radiation Knowledge Label Lentivirus Vector Libraries MLH1 gene Maintenance Malignant Neoplasms Measures Mediating Messenger RNA MicroRNAs Modeling Molecular Nonhomologous DNA End Joining Normal Cell Nucleotide Excision Repair Pathway interactions Phenotype Physiologic pulse Proteins RNA RNA Degradation RNA Stability RNA chemical synthesis Radiation Regimen Role SHFM1 gene Sampling Seeds Single Strand Break Repair Somatic Mutation Stress Techniques Technology Tumor-Derived XPA gene XRCC1 gene base cancer cell chemotherapeutic agent expectation genome-wide homologous recombination improved inhibitor/antagonist neoplastic cell new technology next generation sequencing novel strategies public health relevance repaired response small hairpin RNA stable cell line tool transcriptome sequencing transcriptomics treatment planning tumor vector

项目摘要

DESCRIPTION (provided by applicant): DNA repair pathways maintain the integrity of the genome and it is therefore not surprising that many cancer cells are defective in some aspect of DNA repair. Fortunately, these cancer-specific DNA repair defects offer novel approaches for tumor-selective therapy such as the synthetic lethality of PARP1 inhibitors with BRCA-deficient or homologous recombination (HR)-deficient tumors. An important goal and challenge is thus to identify which tumors are defective in the various specific pathways of DNA repair. It is understood that changes in DNA repair capacity will impact cellular functions both at baseline and in response to genotoxic stress. Further, it has been suggested that these changes in DNA repair capacity may reprogram the cellular transcriptional landscape. Recent evidence suggests that this reprogramming occurs at multiple levels, including changes in the synthesis and stability of specific RNAs, suggesting that DNA repair deficiency may provide unique transcriptional signatures. It is our expectation that the RNA synthesis and stability signature associated with a defect in HR will be unique and may be useful in identifying HR defects without knowledge of any underlying gene defect. We have developed a genome-wide approach for identifying such a signature, termed "BrU-Seq", in which BromoUridine pulse-labeling of nascent RNAs is combined with RNA-Seq in a manner that reveals a remarkably large number of influences on gene expression with a clarity and efficiency not allowed by standard microarray or RNA-Seq approaches. To advance our understanding of the impact of a sudden (somatic) DNA repair defect in a normal cell, we propose to comprehensively evaluate the dynamic status of all coding and regulatory RNAs in the model non-tumorigenic epithelial cell line, MCF-10A, following loss of expression of DNA repair genes across each DNA repair pathway. We hypothesize that DNA repair defects will impact mRNA/microRNA expression and stability with DNA repair pathway-specific signatures that can be used to predict response to certain chemotherapeutic agents. Specifically, we will use BrU-Seq technology to (1) measure the synthesis, stability and structural content of all coding and regulatory RNA species in isogenic MCF-10A cell lines deficient in a key gene in each of the six major DNA repair pathways and (2) evaluate genotoxic stress-induced alterations in the transcriptional profile of the same cells in response to ionizing radiation and the chemotherapeutics temozolimide, cisplatin and PARP inhibitors. These studies will be a critical first step towards identifying signatures associated with specific DNA repair pathway defects in tumor samples that may be used as biomarkers to predict response to targeted chemotherapeutic agents.

描述（由申请人提供）：DNA 修复途径维持基因组的完整性，因此许多癌细胞在 DNA 修复的某些方面存在缺陷也就不足为奇了。幸运的是，这些癌症特异性 DNA 修复缺陷为肿瘤选择性治疗提供了新方法，例如 PARP1 抑制剂对 BRCA 缺陷或同源重组 (HR) 缺陷肿瘤的合成致死作用。因此，一个重要的目标和挑战是确定哪些肿瘤在 DNA 修复的各种特定途径中存在缺陷。据了解，DNA 修复能力的变化将影响细胞的基线功能和对基因毒性应激的反应。此外，有人认为 DNA 修复能力的这些变化可能会重新编程细胞转录景观。最近的证据表明，这种重编程发生在多个层面，包括特定 RNA 合成和稳定性的变化，这表明 DNA 修复缺陷可能提供独特的转录特征。我们期望与 HR 缺陷相关的 RNA 合成和稳定性特征将是独特的，并且可能有助于在不了解任何潜在基因缺陷的情况下识别 HR 缺陷。我们开发了一种全基因组方法来识别这种特征，称为“BrU-Seq”，其中新生 RNA 的溴尿苷脉冲标记与 RNA-Seq 相结合，揭示了对基因表达的大量影响其清晰度和效率是标准微阵列或 RNA-Seq 方法所无法达到的。为了加深我们对正常细胞中突然（体细胞）DNA 修复缺陷的影响的理解，我们建议全面评估模型非致瘤性上皮细胞系 MCF-10A 中所有编码和调节 RNA 的动态状态，如下每个 DNA 修复途径中 DNA 修复基因的表达缺失。我们假设 DNA 修复缺陷将影响 mRNA/microRNA 的表达和 DNA 修复途径特异性特征的稳定性，这些特征可用于预测对某些化疗药物的反应。具体来说，我们将使用 BrU-Seq 技术来 (1) 测量六种主要 DNA 修复途径中关键基因缺陷的同基因 MCF-10A 细胞系中所有编码和调节 RNA 种类的合成、稳定性和结构内容，以及(2) 评估基因毒性应激诱导的相同细胞转录谱的变化，以响应电离辐射和化疗药物替莫唑胺、顺铂和 PARP 抑制剂。这些研究将是识别与肿瘤样本中特定 DNA 修复途径缺陷相关的特征的关键的第一步，这些特征可用作预测对靶向化疗药物的反应的生物标志物。