DNA methylation-based assays for detecting disease spread in rhabdomyosarcoma

基于 DNA 甲基化的检测用于检测横纹肌肉瘤疾病传播

• 批准号: 8100436
• 负责人: STEPHEN R MASTER
• 金额: $ 21.41万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2013-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8100436
• 关键词: Alveolar; Aspirate substance; Biological Assay; Blood Cells; Blood specimen; Bone Marrow; Cells; Characteristics; Childhood; Children' s Oncology Group; Clinical; Clinical Research; Clinical Trials; Collection; Controlled Clinical Trials; Controlled Study; DNA; DNA Methylation; DNA Modification Process; Data; Detection; Diagnosis; Disease; Distal; Failure; Family; Freezing; Frequencies; Gel; Gene Fusion; Genes; Genomics; Goals; Histologic; Hypermethylation; Institution; Investigation; Laboratory Study; Malignant Childhood Neoplasm; Malignant Neoplasms; Methodology; Methods; Methylation; Mutation; Neoplasm Metastasis; Newly Diagnosed; Normal Cell; Oncology Group; Operative Surgical Procedures; Outcome; Patients; Performance; Predictive Value; Primary Neoplasm; Protocols documentation; Publishing; RNA; RNA Degradation; Research Project Grants; Rhabdomyosarcoma; Risk; Risk Assessment; Sampling; Series; Site; Skeletal Muscle; Soft Tissue Neoplasms; Specimen; Staging; System; Testing; Time; Tissues; base; bisulfite; cancer cell; cancer type; design; molecular marker; neoplastic cell; peripheral blood; prognostic; public health relevance; sarcoma; soft tissue; tumor

DESCRIPTION (provided by applicant): Rhabdomyosarcoma (RMS) is a family of pediatric soft tissue tumors related to the skeletal muscle lineage. There have been incremental improvements in outcome over the last three decades, such that cure is now possible for ~70% of newly diagnosed RMS patients. A risk-adapted therapy system has been developed for RMS with risk assessment based on pre-surgical stage, post-surgical group, presence of distal metastases, and histologic subtype. Beyond these clinical indicators of cancer status, molecular markers are now being evaluated to provide further improvement in risk prediction. This application focuses on analysis of minimal disseminated disease in which PCR-based assays of tumor-specific molecular markers permit sensitive detection of tumor cell spread to sites such as bone marrow. For RMS, several small studies used RNA-based PCR methods to demonstrate the feasibility of detecting RMS cells in bone marrow in patients without clinical evidence of metastatic disease. Furthermore, there is evidence from these and other studies that detection of occult tumor cells in the bone marrow is predictive of a worse outcome. To fully understand the utility of this approach in RMS management, these minimal disseminated disease assays must be conducted as part of large well-controlled clinical trials. In an effort to pursue this goal, bone marrow and peripheral blood samples were collected in the recently completed Children's Oncology Group D9803 trial of intermediate risk RMS. Due to the time required for the centralized collection protocol, the RNA in these specimens was often partially degraded. However, the DNA in these samples is still intact and this project will develop DNA-based assays applicable to RMS. DNA hypermethylation is a DNA modification that is often tumor-specific at numerous genomic loci. It can be detected by a PCR-based methodology that has been used to screen for occult cancer cells in multiple cancer types. Based on preliminary evidence that DNA hypermethylation occurs at specific loci in both RMS subtypes, a microarray strategy will be used to screen for genomic loci that are frequently hypermethylated in RMS tumors but not in normal blood cells. These findings will be confirmed and refined by PCR-based assays. To extend these findings, a small panel of quantitative methylation-specific PCR assays will be designed that detect at least one methylation change in most RMS tumors. This panel of quantitative PCR assays will be applied to DNA from the D9803 bone marrow samples as well as a panel of paired primary tumors. The results of these assays will be compared with clinical and pathological characteristics as well as patient outcome to explore the significance of minimal disseminated disease in the bone marrow at diagnosis. This application thus creates a valuable opportunity to identify, develop and test new assays, and then apply these assays to perform correlative laboratory studies on an existing set of annotated biospecimens that are part of an important clinical initiative of the Soft Tissue Sarcoma Committee of the Children's Oncology Group. PUBLIC HEALTH RELEVANCE: This research project directly focuses on the pediatric cancer rhabdomyosarcoma, and proposes an approach to identify patients with early spread of this cancer to distal sites. In this approach, DNA will be isolated from a tissue to which cancer cells often spread, such as bone marrow, and will be screened for the presence of cancer cells by assaying for genetic changes that commonly occur in the cancer but not in the normal version of that tissue. The value of this approach will then be investigated by using these assays on a series of bone marrow samples from a clinical trial of the Children's Oncology Group, and then comparing the assay results with the patients' outcome data.

描述(申请人提供)：横纹肌肉瘤(RMS)是一种与骨骼肌谱系有关的儿科软组织肿瘤家族。在过去的三十年里，预后有了逐渐的改善，现在大约70%的新诊断的RMS患者有可能治愈。风险适应治疗系统已被开发用于RMS，其风险评估基于术前分期、术后分组、是否存在远端转移和组织学亚型。除了这些癌症状态的临床指标外，现在正在评估分子标记，以进一步改进风险预测。这项应用侧重于微小播散性疾病的分析，在这些疾病中，基于肿瘤特异性分子标志物的聚合酶链式反应分析允许灵敏地检测肿瘤细胞扩散到骨髓等部位。对于RMS，几项小型研究使用了基于RNA的PCR方法来证明在没有临床转移疾病证据的患者的骨髓中检测RMS细胞的可行性。此外，从这些和其他研究中有证据表明，在骨髓中检测到隐匿性肿瘤细胞预示着更糟糕的结果。为了充分了解这种方法在RMS管理中的作用，这些最小的播散性疾病检测必须作为大型受控临床试验的一部分进行。为了实现这一目标，在最近完成的儿童肿瘤组D9803中等风险RMS试验中收集了骨髓和外周血样本。由于集中收集协议所需的时间，这些标本中的RNA经常被部分降解。然而，这些样本中的DNA仍然完好无损，该项目将开发适用于RMS的基于DNA的检测方法。DNA高甲基化是一种DNA修饰，在许多基因组座位上往往是肿瘤特异性的。它可以通过一种基于聚合酶链式反应的方法来检测，该方法已被用于在多种癌症类型中筛查隐匿性癌细胞。根据初步证据，DNA高甲基化发生在两种RMS亚型的特定基因座上，微阵列策略将用于筛选在RMS肿瘤中经常发生高甲基化但在正常血细胞中不经常发生的基因组基因座。这些发现将通过基于聚合酶链式反应的分析得到证实和改进。为了扩大这些发现，将设计一小组甲基化特异性的定量PCR分析，以检测大多数RMS肿瘤中至少一个甲基化变化。这一组定量聚合酶链式反应分析将应用于D9803骨髓样本以及一组配对的原发肿瘤的DNA。这些检测的结果将与临床和病理特征以及患者预后进行比较，以探讨骨髓微小播散性疾病在诊断时的意义。因此，这项应用创造了一个宝贵的机会来识别、开发和测试新的分析方法，然后应用这些分析方法对现有的一组带注释的生物标本进行相关的实验室研究，这些生物标本是儿童肿瘤组软组织肉瘤委员会一项重要临床倡议的一部分。 公共卫生相关性：这项研究项目直接关注儿童癌症横纹肌肉瘤，并提出了一种方法来识别这种癌症早期扩散到远端部位的患者。在这种方法中，DNA将从癌细胞经常扩散到的组织中分离出来，如骨髓，并将通过分析在癌症中常见但在正常组织中不出现的基因变化来筛选癌细胞的存在。然后，将通过对儿童肿瘤组临床试验的一系列骨髓样本进行这些分析，然后将分析结果与患者的结果数据进行比较，来研究这种方法的价值。

项目成果

Distinct methylation profiles characterize fusion-positive and fusion-negative rhabdomyosarcoma.

• DOI: 10.1038/modpathol.2015.82
• 发表时间: 2015-09
• 期刊: Modern pathology : an official journal of the United States and Canadian Academy of Pathology, Inc
• 作者: Sun W;Chatterjee B;Wang Y;Stevenson HS;Edelman DC;Meltzer PS;Barr FG
• 通讯作者: Barr FG