A highly sensitive linear amplification based DNA methylation profiling technique for clinical cancer research

用于临床癌症研究的基于高灵敏度线性扩增的 DNA 甲基化分析技术

• 批准号: 10413620
• 负责人: BRIAN C-H CHIU
• 金额: $ 42.12万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-06-08 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10413620
• 关键词: Achievement; Address; Aftercare; Biological Assay; Biological Markers; Biological Process; Biopsy; Blood; Cancer Biology; Cancer Detection; Cancer Diagnostics; Cancer Patient; Cells; Clinic; Clinical; Clinical Oncology; Colon Carcinoma; CpG dinucleotide; Cytosine; DNA; DNA Library; DNA methylation profiling; Detection; Development; Diagnosis; Diagnostic Procedure; Early Diagnosis; Ensure; Environment; Epigenetic Process; FDA approved; Frequencies; Funding; Gene Expression Regulation; Genetic; Genomic DNA; Genomics; Goals; Gold; Guanine + Cytosine Composition; Human; Human Genome; Individual; Investigation; Liquid substance; Machine Learning; Malignant Neoplasms; Malignant neoplasm of liver; Malignant neoplasm of pancreas; Methylation; Mission; Modification; Molecular; Molecular Analysis; Molecular Profiling; Molecular Target; Monitor; Morbidity - disease rate; Multiple Myeloma; Newly Diagnosed; Pathogenesis; Patient Monitoring; Patients; Plasma; Plasma Cells; Population Study; Prevalence; Primary Neoplasm; Procedures; Prognosis; Public Health; RNA; Research; Residual Neoplasm; Resistance; Resolution; Role; Sampling; Screening for cancer; Source; Specificity; Specimen; Systems Biology; Techniques; Technology; Testing; Tissues; Tumor Bank; Tumor Tissue; United States National Institutes of Health; Validation; age group; anticancer research; base; biobank; biomarker discovery; bisulfite; bisulfite sequencing; cancer diagnosis; cancer epidemiology; cancer type; cell free DNA; chemical stability; clinical application; cost efficient; design; epidemiology study; future implementation; high risk population; improved; innovation; innovative technologies; large cell Diffuse non-Hodgkin' s lymphoma; liquid biopsy; minimally invasive; mortality; multidisciplinary; multiple omics; new technology; next generation sequencing; noninvasive diagnosis; novel; prospective; recruit; screening; sex; success; tool; tumor; tumor DNA; tumor heterogeneity; tumor microenvironment; whole genome

PROJECT SUMMARY Validating highly sensitive molecular analysis approaches that can fully exploit precious clinical specimens will transform cancer research and clinical applications. Tissue biopsy is the gold standard for cancer diagnosis and has been the primary source of clinical biospecimens in cancer research. However, one of the biggest challenges in basic and clinical cancer research is the frequent lack of sufficient amount of tumor materials from biopsy for multi-omics research (e.g., parallel RNA and DNA-based profiling) after diagnostic procedures, thus limiting progress in systems biology. As such, the ability to conveniently sample bodily fluids, including circulating cell-free DNA (cfDNA) from plasma, offers great promise for enabling highly-sensitive, minimally- invasive, cost-efficient cancer diagnostic methods, and facilitating screening of high-risk populations and patient monitoring. However, cfDNA typically exists in extremely low quantity (e.g., a few nanograms from several mL of blood). Cancer-derived cfDNA is expected to constitute an even smaller portion of the already scarce cfDNA. To accelerate and enhance cancer biology and clinical applications, this R33 aims to validate a novel technology for profiling DNA methylation, i.e., 5-methylcytosines (5mC), which contributes to cancer pathobiology, is reflected in patient-derived cfDNA, and has been integrated in several FDA-approved tests. Given the prevalence of 5mC in the human genome, its roles in gene regulation, and high chemical stability, validating a novel 5mC technology in nanogram or sub-nanogram-level DNA materials offers promising opportunities for various applications in cancer research that have been limited by technology that can utilize limited clinical samples. Specifically, we developed the T7-Linear Amplification based Bisulfite Sequencing (LABS-seq) that integrates a specially-designed bisulfite conversion procedure with the next-generation sequencing (NGS) for sensitively and unbiasedly detecting 5mC in nanogram or sub-nanogram-level DNA materials (e.g., 100 pg). Our preliminary results demonstrated the technical robustness of the LABS-seq and the feasibility of using this innovative technology to identify cancer-specific 5mC changes in cfDNA. In this R33, we will rigorously validate the LABS-seq technique using banked tumor tissues/cells and plasma cfDNA samples from 250 patients with diverse cancer types and 50 frequency-matched healthy controls as well as longitudinal samples from 200 prospectively recruited cancer patients. In Aim 1, we will validate the LABS-seq in genomic DNA (gDNA) from tumor tissues/cells to detect cancer type-specific epigenetic alterations. In Aim 2, we will validate the LABS-seq in cfDNA for the detection of cancer and longitudinal changes. Upon completion of this project, we will provide a highly sensitive, cost-efficient, transformative epigenetic approach applicable to both gDNA and cfDNA, opening up opportunities for research that have been limited by technology. Our established multidisciplinary team, well-annotated human clinical specimens, and excellent environment will ensure the success of this proposed project and future implementation in the clinic.

项目摘要 验证可以完全利用宝贵临床标本的高度敏感的分子分析方法 改变癌症研究和临床应用。组织活检是癌症诊断的黄金标准 并一直是癌症研究中临床生物测量的主要来源。但是，最大的 基本和临床癌症研究中的挑战是经常缺乏足够数量的肿瘤材料 诊断程序之后的多词研究活检（例如，平行RNA和基于DNA的分析）， 因此限制了系统生物学的进展。因此，可以方便地采样体液的能力，包括 血浆中循环无细胞的DNA（CFDNA）为实现高度敏感的，最小的 - 侵入性，具有成本效益的癌症诊断方法，并促进筛查高危人群和 患者监测。但是，CfDNA通常以极低的数量存在（例如，来自 几毫升的血液）。癌症衍生的CFDNA有望构成已经较小的已经 稀缺的cfDNA。为了加速和增强癌症的生物学和临床应用，该R33旨在验证 用于分析DNA甲基化的新型技术，即5-甲基环氨酸（5MC），这有助于癌症 病理生物学，反映在患者衍生的CFDNA中，并已集成在几个FDA批准的测试中。 鉴于人类基因组中5MC的流行，其在基因调节中的作用和高化学稳定性， 验证新颖的5MC技术中的纳米图或亚纳米图级DNA材料提供了有希望的 可以利用可以利用的技术限制癌症研究中各种应用的机会 有限的临床样品。具体而言，我们开发了基于T7线性扩增的亚硫酸盐测序 （Labs-seq）将特殊设计的Bisulfite转换过程与下一代集成 测序（NGS），用于敏感和公正地检测纳米图或亚纳图级DNA中的5MC 材料（例如100 pg）。我们的初步结果证明了实验室seq的技术鲁棒性和 使用这种创新技术鉴定CFDNA中癌症特异性5MC变化的可行性。在这个R33中， 我们将使用银行肿瘤组织/细胞和等离子体CFDNA严格验证实验室-Seq技术 来自250例癌症类型和50个频率匹配的健康对照的患者的样本以及 来自200名前瞻性招募的癌症患者的纵向样本。在AIM 1中，我们将验证Labs-seq 在肿瘤组织/细胞的基因组DNA（GDNA）中，以检测癌症型特异性表观遗传学改变。在AIM 2中， 我们将在CFDNA中验证实验室seq，以检测癌症和纵向变化。完成后 在这个项目中，我们将提供一种适用的高度敏感，成本效益，变革性的表观遗传方法 对于GDNA和CFDNA，开放了受技术限制的研究机会。我们的 建立的多学科团队，通知良好的人类临床标本和出色的环境将会 确保该提议的项目和未来在诊所实施的成功。