Validation of copy number changes by MLPA as predictors of relapse in Wilms tumor

验证 MLPA 的拷贝数变化作为肾母细胞瘤复发的预测因素

• 批准号: 8330806
• 负责人: Elizabeth J Perlman
• 金额: $ 15.99万
• 依托单位: LURIE CHILDREN'S HOSPITAL OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-09 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8330806
• 关键词: 15q; 16q; 17p; 22q; Biological Assay; Biological Markers; Child; Childhood; Children' s Oncology Group; Chromosomal translocation; Chromosomes; Chromosomes, Human, Pair 1; Clinical; Constitutional; Cytogenetic Analysis; DNA; Data; Future; Gene Expression; Genes; Goals; Histology; Isochromosomes; Laboratories; Ligation; Loss of Heterozygosity; Malignant neoplasm of kidney; Measures; Methylation; Microsatellite Repeats; Mutation; National Wilms' Tumor Study Group; Nephroblastoma; Outcome; Patients; Prevalence; Protocols documentation; Reagent; Recurrence; Relapse; Relative Risks; Reporting; Resources; Risk; Sampling; Staging; Technology; Testing; Tissues; Toxic effect; Unbalanced Translocation; Uniparental Isodisomies; Validation; adverse outcome; arm; chemotherapy; chromosome 1q gain; cohort; comparative genomic hybridization; cost; design; flexibility; follow-up; high risk; imprint; improved; reagent testing; research clinical testing; transcriptomics; tumor

DESCRIPTION (provided by applicant): Favorable Histology Wilms Tumor (FHWT) is the most common malignant renal tumor of childhood. Approximately 15% of FHWT relapse, and of these only 50% survive. Currently, loss of heterozygosity (LOH) for chromosomes 1p and 16q have been demonstrated and validated to be associated with poor outcome for patients with FHWT. Children registered on the current Children's Oncology Group (COG) protocols are now stratified for therapy according to their 1p and 16q LOH status. However, both 1p and 16q LOH is present in only 5% of FHWT, and predicts only 9% of relapses. Additional markers for relapse are needed. Recently, multiple studies using multiple different technologies have independently concluded that 1q gain (identified in approximately 25% of FHWT) is a much stronger predictor of relapse in FHWT than LOH of either 1p or 16q. Furthermore, in some patients these markers are inter-related due to the presence of unbalanced chromosomal translocations. Alterations to several other loci have also been identified in FHWT due to their association with relapse or acquisition following relapse (15q gain, 17p loss, and 22q loss). These changes have not been tested and validated in prospectively identified patients. This study seeks to develop and validate an inexpensive, robust clinical assay that may be utilized to stratify patients in the next COG protocol. Multiplex Ligation-dependent Probe Amplification (MLPA), was chosen due its simplicity, flexibility, sensitivity, amenability to multiplexing, applicability to archival tissues, and availability within clinical laboratories. Aims 1-3: To analyze 250 FHWT registered on NWTS-4 for 1q, 1p, 16q, 15q, 17p, and 22 copy number using MLPA. Samples previously analyzed for 1p and 16q LOH will be studied using a commercially available MLPA test that detects copy number of subtelomeric loci on each chromosomal arm (46 total targets). Chromosomal regions significantly associated with survival will be identified, and multiple probes will be designed for each chromosomal region identified and these will be multiplexed into a single clinical test. Aim 4: To validate the targeted multiplex MLPA test developed in Aims 1-3 on an independent set of 600 FHWT registered on NWTS-5. A case-cohort of 600 patients enriched for relapse has been developed from NWTS-5. The association between copy number of each locus and survival will be determined. Relevance: This study will result in a robust clinical test that requires fewer resources and will result in more information than is currently available. The currently performed microsatellite analysis for 1p and 16q LOH costs ~$50/test for reagents alone and requires both constitutional and tumor DNA. A targeted MLPA test will cost only $5/test, and will require only tumor DNA. The final MLPA test will analyze a minimum of three loci with multiple targets per locus and will be capable of measuring 100 targets. With a conservative projected relative risk of 2.0 and a prevalence of 25%, a minimum of 40% of relapses will be predicted. If so validated, the MLPA assay will be used to stratify therapy in the COG protocol that is anticipated to open in 2013.

肾母细胞瘤（Wilms Tumor， FHWT）是儿童时期最常见的恶性肾脏肿瘤。大约15%的FHWT复发，其中只有50%存活。目前，染色体1p和16q的杂合性缺失（LOH）已被证实与FHWT患者预后不良相关。在目前的儿童肿瘤组（COG）协议中注册的儿童现在根据他们的1p和16q LOH状态进行分层治疗。然而，1p和16q LOH仅出现在5%的FHWT中，并且仅预测9%的复发。需要其他复发标志物。最近，使用多种不同技术的多项研究独立得出结论，1q增加（约25%的FHWT）是FHWT复发的更强预测因子，而不是1p或16q的LOH。此外，在一些患者中，由于存在不平衡的染色体易位，这些标记是相互关联的。由于FHWT与复发或复发后获得相关（15q增加，17p损失和22q损失），在FHWT中也发现了其他几个位点的改变。这些变化尚未在前瞻性确定的患者中进行测试和验证。本研究旨在开发和验证一种廉价、可靠的临床检测方法，可用于在下一个COG方案中对患者进行分层。选择多重连接依赖探针扩增（MLPA），是因为其简单、灵活、敏感、可复用、适用于档案组织和临床实验室的可用性。目标1-3：使用MLPA分析在NWTS-4上注册的250个FHWT的1q、1p、16q、15q、17p和22个拷贝数。先前分析的样品1p和16q LOH将使用市售的MLPA测试进行研究，该测试检测每条染色体臂上亚端粒位点的拷贝数（总共46个靶标）。将确定与生存显著相关的染色体区域，并为每个已确定的染色体区域设计多个探针，这些探针将多路复用到单个临床测试中。目标4：在NWTS-5上注册的600台独立FHWT上验证目标1-3中开发的靶向多路MLPA测试。从NWTS-5开发了一个600例复发患者的病例队列。将确定每个基因座的拷贝数与存活率之间的关系。相关性：这项研究将产生一个强有力的临床试验，所需的资源更少，并将产生比目前可用的更多的信息。目前进行的微卫星分析1p和16q LOH的成本约为50美元/单试剂，并且需要体质和肿瘤DNA。一个有针对性的MLPA测试只需要5美元/次，并且只需要肿瘤DNA。最终的MLPA测试将分析至少三个基因座，每个基因座有多个目标，并将能够测量100个目标。保守估计相对危险度为2.0，患病率为25%，预计至少有40%的复发。如果得到验证，MLPA分析将在COG方案中用于分层治疗，预计将于2013年开放。