Author: Guanhua Zhu, Assistant Professor (a, b, c); Peiyong Jiang, Associate Professor (a, b, c, d)
a. Centre for Novostics, Hong Kong Science Park, Pak Shek Kok, Hong Kong SAR, China
b. Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Shatin, Hong Kong SAR, China
c. Department of Chemical Pathology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, Hong Kong SAR, China
d. State Key Laboratory of Translational Oncology, The Chinese University of Hong Kong, Prince of Wales Hospital, Shatin, Hong Kong SAR, China
Editor: Jada Yu Zhang, Assistant Professor, UT MD Anderson Cancer Center
Cell-free DNA (cfDNA) refers to fragmented DNA molecules circulating in the bloodstream, released from dying cells in both physiological and pathological process. The analysis of cfDNA through liquid biopsy—a minimally invasive approach that detects molecular biomarkers from blood samples—has gained enormous interest in recent years. cfDNA-based biomarkers offer promising applications in early cancer detection, disease monitoring, and prenatal testing. Among various cfDNA characteristics, DNA methylation has attracted particular attention due to its role in gene regulation and tissue-specific patterns. Methylation-associated fragmentation signatures provide a unique opportunity to enhance the sensitivity and specificity of liquid biopsy-based diagnostics by integrating epigenetic and fragmentomic information.
Dr. Guanhua Zhu and Dr. Peiyong Jiang are from Dr. Y.M. Dennis Lo’s team at the Chinese University of Hong Kong. The team pioneers in cfDNA research and clinical applications, have made significant contributions to the field. In 2022, their group reported a study in PNAS introducing Fragmentomics-based Methylation Analysis (FRAGMA), a novel approach for analyzing short-range cfDNA cleavage patterns associated with cytosine-phosphate-guanine (CpG) methylation (1). This technique provided insights into the epigenetic regulation of cfDNA and demonstrated its feasibility in distinguishing between healthy and diseased states, particularly in oncology and prenatal diagnostics.
Building upon this foundation, the team further expanded their work in a 2024 Clinical Chemistry study, introducing FRAGMAXR, an extension of FRAGMA that examines cfDNA nucleosomal patterns over longer genomic distances, spanning multiple nucleosomes (2). This study demonstrated that FRAGMAXR effectively detects cancer, with a strong correlation between its signals and hepatocellular carcinoma (HCC) stages. FRAGMAXR achieved an area under the receiver operating characteristic curve (AUC) of 0.93 for distinguishing HCC patients from non-cancer individuals. Additionally, it demonstrated stage-dependent sensitivity, detecting 58% of early-stage, 89% of intermediate-stage, and 95% of advanced-stage HCC patients at 99% specificity. Not limited to HCC, the method was also applied to multicancer detection, achieving AUC values of 0.99 for lung cancer, 0.81 for breast cancer, and 0.93 for ovarian cancer. The nucleosomal patterns-based approach for cancer detection can be generalized across different datasets. Moreover, the combination of FRAGMA and FRAGMAXR further enhanced diagnostic performance, improving the AUC to 0.98 in HCC while reducing false negatives.
Given its ability to identify tissues of origin, FRAGMAXR has significant implications beyond oncology. Its potential applications extend to noninvasive prenatal testing, where accurate tissue-of-origin analysis is crucial. FRAGMAXR has demonstrated high accuracy in estimating fetal DNA contributions. The findings from this study mark an important advancement in cfDNA-based diagnostics, offering a powerful tool for early disease detection and personalized medicine.
Figure. Schematic illustration for fragmentomics-based methylation analysis of cell-free DNA, including FRAGMA (left) and FRAGMAXR (right).
游离DNA片段组学的新前沿：甲基化相关的核小体DNA断裂模式
游离DNA（cfDNA）是指循环于血液中的片段化 DNA 分子，由细胞在生理或病理死亡过程中释放而来。近年来，通过液体活检对 cfDNA 进行分析——这是一种从血液样本中检测分子生物标志物的微创方法——已引起广泛关注。 cfDNA 作为标志物在癌症早期检测、疾病监测和产前筛查方面具有广阔的应用前景。在cfDNA 的众多特征中，DNA 甲基化因其在基因调控中的重要作用及组织特异性而备受关注。与甲基化关联的片段化特征通过整合表观遗传学和片段组学信息，为液体活检诊断提供了独特机会，提高检测的敏感性和特异性。
朱冠华教授和江培勇教授来自香港中文大学的卢煜明教授团队。该团队是 cfDNA 研究及临床应用领域的先驱，他们在该领域做出了重要贡献。2022 年，他们在 PNAS 发表研究，提出基于片段组学的甲基化分析（FRAGMA）(1)。 作为一种新颖的方法，FRAGMA可用于分析与胞嘧啶-磷酸-鸟嘌呤（CpG）甲基化相关的短程 cfDNA 断裂模式。该技术为 cfDNA 的表观遗传调控提供了新的见解，并验证了其区分健康与疾病状态的可行性，特别是在肿瘤学和产前诊断领域。
在此基础上，该团队在 2024 年 Clinical Chemistry 研究中进一步拓展了他们的工作，研发出FRAGMAXR，可用于分析较长基因组距离范围内的 cfDNA 核小体模式(2)。研究表明，FRAGMAXR 能够有效检测癌症，其信号与肝细胞癌（HCC）的密切相关。该方法在区分 HCC 患者与非癌症个体方面，受试者工作特征曲线下面积（AUC）达 0.93。此外，它表现出分期依赖的敏感性，在 99% 的特异性下，检测出 58% 的早期 HCC 病例，89% 的中期病例，95% 的晚期病例。FRAGMAXR 不仅限于 HCC，还可用于多癌种检测，在肺癌、乳腺癌和卵巢癌中的 AUC 分别达到 0.99、0.81 和 0.93。该基于核小体模式的癌症检测方法可推广至不同的数据集。此外，FRAGMA 与 FRAGMAXR 的结合进一步提高了对肝细胞癌的诊断性能，使 AUC 提升至 0.98，并降低了假阴性率。
由于 FRAGMAXR 能够识别组织来源，其意义不仅局限于肿瘤学，还具有更广泛的应用前景。例如，它可用于无创产前筛查，其中精准的组织来源分析至关重要。FRAGMAXR在估算胎儿DNA贡献方面表现出高度准确性。本研究的发现代表了 cfDNA 诊断领域的一项重要进展，为早期疾病检测和个性化医疗提供了强有力的工具。
References
1. Zhou Q, Kang G, Jiang P, Qiao R, Lam WKJ, Yu SCY, et al. Epigenetic analysis of cell-free DNA by fragmentomic profiling. Proc Natl Acad Sci U S A 2022 Nov;119:44:e2209852119. Epub 20221026 as doi: 10.1073/pnas.2209852119. https://doi.org/10.1073/pnas.2209852119
2. Zhu G, Jiang P, Li X, Peng W, Choy LYL, Yu SCY, et al. Methylation-Associated Nucleosomal Patterns of Cell-Free DNA in Cancer Patients and Pregnant Women. Clin Chem 2024 Nov 4;70:11:1355-65 as doi: 10.1093/clinchem/hvae118. https://doi.org/10.1093/clinchem/hvae118
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