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: 7875543
• 负责人: FREDERIC G BARR
• 金额: $ 17.4万
• 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 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7875543
• 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（由申请人提供）：横纹肌肉瘤（RMS）是与骨骼肌谱系有关的小儿软组织肿瘤系列。在过去的三十年中，结果已经有所改善，因此，对于新诊断的RMS患者，现在可以治愈可能治愈。已经为RMS开发了一种风险适应的治疗系统，该RMS具有基于手术前阶段，手术后组，远端转移和组织学亚型的风险评估。除了这些癌症状态的临床指标之外，现在还对分子标记物进行评估，以进一步改善风险预测。该应用集中在分析最小传播疾病的分析中，其中基于PCR的肿瘤特异性分子标记物的测定允许对肿瘤细胞扩散到骨髓等部位的敏感检测。对于RMS，几项小型研究使用基于RNA的PCR方法来证明在没有转移性疾病的临床证据的患者中检测骨髓中RMS细胞的可行性。此外，从这些和其他研究中有证据表明，骨髓中隐匿性肿瘤细胞的检测可预测结果较差。为了充分了解这种方法在RMS管理中的实用性，这些最少的传播疾病测定必须作为大型控制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学组D9803中级风险RMS试验中收集了骨髓和外周血样本。由于集中式收集方案所需的时间，这些标本中的RNA通常会部分降解。但是，这些样品中的DNA仍然完整，该项目将开发适用于RMS的基于DNA的测定。 DNA高甲基化是一种DNA修饰，在许多基因组基因局中通常是肿瘤特异性的。可以通过基于PCR的方法来检测它，该方法已用于筛选多种癌症类型的隐匿性癌细胞。基于初步证据表明，DNA高甲基化发生在两个RMS亚型中的特定基因座上，将使用微阵列策略用于筛选基因组基因群，这些基因组基因群经常在RMS肿瘤中经常被过度甲基化，但在正常血细胞中却没有。这些发现将通过基于PCR的测定法确认和完善。为了扩展这些发现，将设计一小部分定量甲基化特异性PCR测定法，以检测大多数RMS肿瘤中至少一种甲基化变化。该定量PCR分析将用于D9803骨髓样品以及一组配对的原发性肿瘤的DNA。这些测定的结果将与临床和病理特征以及患者结局进行比较，以探讨诊断时骨髓中最小传播疾病的重要性。因此，该应用创造了一个宝贵的机会，可以识别，开发和测试新的测定法，然后对这些测定法进行相关的实验室研究，以对现有的带注释的生物素质群进行相关的实验室研究，这是儿童肿瘤学小组软组织肉瘤委员会一项重要临床计划的一部分。 公共卫生相关性：该研究项目直接关注儿科癌症肉瘤，并提出了一种方法来鉴定该癌症早期扩散到远端部位的患者。在这种方法中，将从癌细胞经常扩散的组织中分离出DNA，例如骨髓，并通过分析通常在癌症中发生但不在正常版本中的遗传变化来筛选癌细胞的存在。然后，将通过在儿童肿瘤学组的临床试验的一系列骨髓样品中使用这些测定方法来研究这种方法的价值，然后将测定结果与患者的结果数据进行比较。