Comprehensive minimal residual disease tracking in cancer

癌症的全面微小残留病追踪

• 批准号: 9920128
• 负责人: G. Mike Makrigiorgos
• 金额: $ 37.96万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-06-01 至 2023-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9920128
• 关键词: Admixture; Alleles; Bar Codes; Biological Assay; Biopsy; Blood; Cancer Patient; Cells; Clinical; Clinical Research; Clinical Sensitivity; Clinical Trials; Computing Methodologies; Consensus Sequence; DNA; Data; Detection; Detection of Minimal Residual Disease; Disease; Early Diagnosis; Fingerprint; Genes; Health Care Costs; Image; Intervention; Local Therapy; Malignant Neoplasms; Methods; Microscopic; Molecular; Molecular Profiling; Monitor; Morbidity - disease rate; Mutation; Mutation Detection; Names; Noise; Patient observation; Patients; Plasma; Primary Neoplasm; Probability; Public Health; Relapse; Residual Cancers; Residual Neoplasm; Residual Tumors; Sampling; Sensitivity and Specificity; Systemic Therapy; Technology; Testing; Time; Tumor Markers; Tumor-Derived; Variant; base; cancer therapy; cell free DNA; circulating DNA; cost effective; digital; exome sequencing; follow-up; individual patient; melanoma; mutant; next generation sequencing; nuclease; overtreatment; prevent; prognostic value; screening; side effect; tumor

Identifying the right amount of therapy – no more and no less – for patients with early stage cancer remains a challenge because there is no reliable method by which to separate those with microscopic residual disease after systemic or local therapies, from those without it. Current imaging can barely detect a mass of 1 million cells, while it takes just one cell to spawn new tumors that, by the time they are detected, are often incurable. Early detection of minimal residual disease (MRD) could give patients who need further treatment a chance at a cure, and prevent over-treatment of others. Despite its promise, MRD detection based on technologies like digital PCR that detect a single tumor marker at a time has inadequate ability to detect residual cancer at early stages. Next generation sequencing (NGS) can track many mutations simultaneously; however NGS requires extensive corrections using molecular barcoding to reduce noise and detect low-level mutations. This requirement invariably diminishes NGS throughput and increases expense. Currently, for NGS it is either sequencing depth or breadth, but not both. Here we propose to refine and apply a transformative technology that enables highly sensitive tracing of MRD in blood despite limited cfDNA material, while also retaining NGS throughput (breadth) and depth. We recently developed NaME-PrO, a simple and powerful technology that enables NGS to track extremely low-level mutations in circulating DNA. NaME-PrO utilizes a nuclease guided by probes to thousands of DNA targets, to render WT sequences non-amplifiable thereby allowing mutation– containing sequences to amplify and be sequenced with few reads as if they were high abundance mutations. To track MRD in blood, we first create a tumor fingerprint for each patient using whole exome sequencing of the primary tumor to define 30-100 tumor-specific clonal mutations and encompassing truncal mutations. These will be tracked in cfDNA using NaME-PrO-enhanced NGS. NaME-PrO will be combined with molecular barcoding (qNaME-PrO) to enable quantification of the original mutation fraction with few sequencing reads for the patient-specific mutations tracked and elimination of errors. We will (a) optimize and test the use of molecular barcoding in conjunction with NaME-PrO mutation enrichment for quantification of original mutation abundance; (b) Perform exome sequencing to derive mutational tumor fingerprints; then follow fingerprints in plasma and serial dilutions in WT plasma to determine the lowest limit of quantitative detection; and (c) perform a preliminary assessment of the prognostic ability of MRD in melanoma patients. If the project is successful, it will be followed by practice- changing clinical studies. The proposed method is anticipated to provide a high negative predictive power, as one of the main advantages. This could eventually enable `watchful waiting' strategies for some patients currently treated unnecessarily, thus reducing morbidity and health care costs.

对于早期癌症的患者，确定适量的治疗方法（不再和较少） 仍然是一个挑战，因为没有可靠的方法可以将显微镜分开 全身或局部疗法后的残留疾病，来自没有疾病。当前的成像几乎无法检测到 质量为100万个细胞，而仅需一个细胞就产生新的肿瘤，这些肿瘤被发现时， 通常是无法治愈的。早期发现最小残留疾病（MRD）可以为需要进一步的患者提供 治疗有机会治愈的机会，并防止他人过度治疗。尽管有承诺，MRD检测 基于像数字PCR这样的技术，一次检测单个肿瘤标记的能力不足 在早期检测残留癌。下一代测序（NGS）可以跟踪许多突变 相似地;但是，NGS需要使用分子条形码来降低噪声的大量校正 并检测低级突变。此要求总是减少NGS吞吐量并增加 费用。目前，对于NGS，它是测序深度或广度，但并非两者兼而有之。 在这里，我们建议完善和应用一种变革性技术，以使高度敏感 在血液目的地限制cfDNA材料中的MRD跟踪，同时还保留NGS吞吐量（广度） 和深度。我们最近开发了Name-Pro，这是一种简单而强大的技术，使NGS能够 在循环DNA中跟踪极低的突变。名称 - pro利用了问题指导的核酸酶 到成千上万的DNA靶标，使WT序列不可放大，从而使突变 - 包含序列以放大和测序几乎没有读取，好像它们是高丰度 突变。要追踪血液中的MRD，我们首先使用整个外显子为每个患者创建肿瘤指纹 原发性肿瘤的测序以定义30-100个肿瘤特异性克隆突变并包含 截短突变。这些将使用名称增强的NG在CFDNA中跟踪。名称将是 与分子条形码（Qname-Pro）结合起来，以实现原始突变 对于跟踪的患者特异性突变和消除错误的患者特异性突变的分数很少。 我们将（a）优化和测试分子条形码与名称-Pro突变的使用 量化原始突变丰度的富集； （b）执行外显子组测序以得出 突变肿瘤指纹；然后在WT等离子体中遵循血浆中的指纹和连续稀释液 确定定量检测的最低限制； （c）对 MRD在黑色素瘤患者中的预后能力。如果该项目成功，则将进行实践 - 改变临床研究。预计所提出的方法将提供高阴性预测 权力，作为主要优势之一。这最终可以实现“注意等待”策略 一些患者目前对不必要的治疗，从而降低了发病率和医疗费用。