Establishment of a novel genomic approach to non-invasive therapeutic response assessment & monitoring of minimal residual disease (MRD) in patients with Non-Hodgkin´s Lymphoma

我们建议开发和应用一种灵敏的检测方法，用于非霍奇金淋巴瘤（NHL）患者的反应评估，包括MRD（微小残留病）检测，使用一种新的方法定量循环中的肿瘤衍生DNA。循环肿瘤DNA代表了潜在的液体活检，这是一种有前途的肿瘤动力学微创监测生物标志物，因为肿瘤DNA不断被癌细胞释放到血液中，并且可以作为无细胞DNA（cfDNA）检测。高通量测序（HTS）技术的进步已经实现了癌症全基因组的快速测序，并揭示了大多数肿瘤基因组（包括NHL）包含数千种以前未被发现的畸变，这些畸变可以作为NHL患者的生物标志物。通过分析NHL中复发的体细胞畸变的选择性但广泛的集合，我们希望设计一种敏感的早期反应评估和MRD检测方法，用于未来的临床研究和临床使用。使用HTS定量循环肿瘤DNA的主要障碍是必须检测的肿瘤来源的cfDNA的相对低的水平，需要极高的（并且因此昂贵的）测序深度。为了克服这个问题，设计了一种称为CAPP-Seq（CAPP-Seq）的新方法。CAPP-Seq利用定制设计的基于溶液的杂交方法来捕获高度富集单核苷酸变体（SNV）、插入/缺失（indel）或NHL中的易位的基因组区域。本研究的目的是充分开发和表征一种新的下一代测序（NGS）方法，用于检测血液中循环的肿瘤来源的无细胞DNA。我们将使用HTS研究的>230种不同淋巴恶性肿瘤的集合来扩展CAPP-Seq。此外，我们希望通过前瞻性地建立灵敏度和特异性的基准来比较CAPP-Seq与IgH-HTS和标准化放射学标准在NHL中用于NHL应答评估的潜在效用。

We propose to develop and apply a sensitive assay for response assessment including MRD (minimal residual disease) detection in patients with Non-Hodgkins Lymphomas (NHL), using a novel method for quantitation of tumor-derived DNA in circulation. Circulating tumor DNA represents a potential liquid biopsy, a promising biomarker for minimally invasive monitoring of tumor dynamics, since tumor DNA is continually released into the blood by cancer cells and can be detected as cell-free DNA (cfDNA). Advances in high throughput sequencing (HTS) technologies have enabled rapid sequencing of cancer whole genomes and revealed that most tumor genomes, including NHLs, contain thousands of previously unappreciated aberrations, which could serve as biomarkers in NHL patients. Through profiling of a select yet broad collection of somatic aberrations that are recurrent in NHL, we want to devise a sensitive method for early response assessment and MRD detection for future clinical studies and clinical use. A major hurdle in quantitating circulating tumor DNA using HTS is the relatively low level of tumor-derived cfDNA that must be detected, requiring extremely high (and therefore costly) sequencing depth. To overcome this problem a novel approach called CAncer Personalized Profiling by Deep Sequencing (CAPP-Seq) was devised. CAPP-Seq utilizes a custom designed solution-based hybridization method to capture genomic regions highly enriched for single nucleotide variants (SNVs), insertions/deletions (indels), or translocations in NHLs. Aim of this study is to fully develop and characterize a novel next generation sequencing (NGS) method for detecting tumor-derived cell-free DNA circulating in blood. We will expand CAPP-Seq using a collection of >230 diverse lymphoid malignancies studied by HTS. Furthermore, we want to compare the potential utility of CAPP-Seq with IgH-HTS and standardized radiographic criteria for NHL response assessment in NHL by prospectively establishing benchmarks for sensitivity and specificity.